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COLLOIDS  IN  BIOLOGY 
AND  MEDICINE 


BY 

PROF.  H.  BECHHOLD 

Memher  of  the  Royal  Institute  for  Experimental  Therapeutics  in  FranJ:fort  A.  M. 

AUTHORIZED   TRAXSLATIOX   FROM   THE   SECOND   GERMAN 
EDITION,    AVITH   NOTES   AND   EMENDATIONS 

BY 

JESSE   G.  M.  BULLOSA,  A.B..  M.D. 

Assistant  Clinical  Professor   of  Medicine,  Fordham    University,   Adjunct  Professor  of 

Clinical  Medicine,  Xeic  York  Polyclinic  School  and  Hospital.  Visiting  Physician 

Riverside  Sanatorium,  Associate  Visiting  Physician,  Willard 

Parker  Hospital,  New  York  City. 


54   ILLZ'STBATIOXS 


NETV    YORK 

D.    VAX    NOSTRAXD    COMPANY 

2ö  Park  Place 

1919 


Copyright,  1919, 

BY 

D.  VAN  NOSTRAND   COMPANY 


B3g 


StanlJopc  iprcss 

F.    H.GILSON    COMPANY 
BOSTON,  U.S.A. 


AUTHOR'S  .  PREFACE. 

This  book  is  an  attempt  to  apply  the  results  of  colloid  research 
to  biology.  The  reader  may  find  the  undertaking  somewhat  bold, 
since  the  number  of  facts  are  so  few,  and  the  gaps  so  numerous  that 
a  complete  picture  is  impossible.  I  find  myself  somewhat  in  the 
position  of  a  palaeontologist  who  wishes  to  reconstruct  the  ancestry 
of  the  entire  organized  world  from  some  chance  fragments.  Each 
day  brings  new  finds  which  must  be  fitted  into  his  plan  and  which 
confirm  his  views  or  show  that  he  has  been  on  a  false  trail.  From 
the  nature  of  things,  it  happens  that  I  must  more  often  indicate 
problems  than  report  experimental  results.  This  probably  will 
prove  a  stimulus  to  those  who  wish  to  take  active  part  in  the  de- 
velopment of  our  young  science. 

I  wish  to  state  one  additional  fact:  it  was  not  my  purpose  to 
make  this  book  exhaustive.  I  have  endeavored  to  give  a  general 
view,  and  since  the  work  is  addressed  to  biologists  and  physicians  as 
well  as  colloid  investigators,  I  have  striven  to  give  a  clear  picture  of 
the  subject,  disregarding  moot  questions.  —  On  this  account  the 
"Introduction  to  The  Study  of  Colloids"  was  abbreviated  as  much  as 
possible  without  danger  of  obscuring  the  subsequent  parts.  —  Those 
who  wish  to  stud}^  more  thoroughly  pure  colloid  chemistry,  I  refer 
to  the  excellent  books  of  Herbert  Freundlich  "Kapillarchemie" 
Leipzig,  1909,  and  Wolfgang  Oswald  "Grundris  der  Kolloidchemie," 
Dresden. 

Accordingly,  in  the  arrangement  of  the  first  part,  I  have  not  fol- 
lowed the  usual  system,  but  have  been  guided  by  a  desire  to  make 
easy  of  comprehension  the  matters  most  important  to  biologists  and 
physicians.  On  this  account  I  have  considered  it  advisable  to  de- 
vote considerable  space  to  the  "Methods  of  Colloid  Research." 

Some  new  unpublished  experimental  data  of  my  own  and  some 
placed  at  my  disposal  by  others  have  been  included. 

It  is  finally  my  privilege  to  thank  all  those  who  have  helped  me 
in  the  preparation  of  this  book,  particularly  Professors  H.  Apolant, 
R.  Höber,  H.  Sachs,  and  Dr.  H.  Siedentopf.  I  am  especially  in- 
debted to  my  dear  friend,  Professor  Richard  Lorenz,  with  whom  I 
have  discussed  some  of  the  chapters,  and  to  Dr.  and  Mrs.  Ziegler, 


iv  PREFACE 

who  undertook  not  only  the  laborious  task  of  reading  the  proofs  but 
also  checked  back  the  index,  page  for  page.  I  am  also  grateful  to 
my  publisher,  Mr.  Theodor  Steinkopff,  who,  by  intelligent  coopera- 
tion was  of  great  assistance  to  me. 

H.   BECHHOLD. 
Frankfort  on  the  Main 


DEDICATED 

TO  HIS   EXCELLENCY 

PRIVY  MEDICAL  COUNCILLOR 

PROF.  DK   PAUL  EHRLICH 

AND 

PRIVY  SANITARY  COUNCILLOR 

DR.  THEODORE  NEÜBÜRGER 
WITH  THE  GRATITUDE  OF  THE  AUTHOR 


TRANSLATOR'S   PREFACE 

The  translation  of  the  first  edition  of  this  book  was  in  hand  when 
Professor  Bechhold  announced  the  preparation  of  a  second  edition. 
The  translation  was  therefore  delayed  awaiting  that  issue  in  order  to 
bring  the  volume  up  to  date,  and  embody  data  obtainable  from 
recent  literature. 

The  proof  sheets  were  received  in  1915  and  1916,  but  the  transla- 
tion and  revision  were  again  delayed  by  occupation  connected  with  the 
prosecution  of  the  War.     (Local  Board  Membership.) 

The  translator  hopes  that  Professor  Bechhold's  presentation  of  the 
colloid  chemical  problems  of  biology  and  medicine  will  serve  to  stim- 
ulate greater  interest  in  colloid  chemistry  among  physicians,  biolo- 
gists, and  bio-chemists.  His  own  reward,  he  has  already  found  in 
the  interest  attached  to  interpreting  the  every-day  problems  of  prac- 
tice from  this  angle. 

It  is  to  be  expected  that  the  application  of  colloid  principles  to  the 
phenomena  of  life  will  lead  to  new  and  more  rational  theories  of  their 
disturbances,  and  it  is  hoped  that  an  improved  practice  of  medicine 
will  eventually  result  The  pages  of  this  book  indicate  the  begin- 
nings already  made  and  suggest  future  studies. 

The  translator  acknowledges  his  debt  to  Dr.  B.  Michaelovsky  for 
assistance  with  the  translation;  to  Mrs.  D.  D.  Pool  for  the  author 
index,  and  to  his  publishers,  Messrs.  D.  Van  Nostrand  Company,  for 
their  friendly  co-operation. 

His  debts  to  his  friend,  Jerome  Alexander,  he  is  unable  to  enu- 
merate. He  owes  to  him,  not  only  his  introduction  to  colloid  chem- 
istry, but  never  failing  stimulation,  encouragement  and  assistance 
with  the  translations  and  the  proof-reading:  Any  merit  the  work 
possesses  is  due  to  his  collaboration. 

J.  G.  M.  B. 
February  15,  1919. 
62  West  Eighty-Seventh  Street, 
New  York  City. 


THIS  TRANSLATION   IS 

DEDICATED   WITH  AFFECTIONATE  GRATITUDE 

TO    MY    SISTER 

EMILIE  M.  BULLOWA,  L.L.B 


TABLE  OF   CONTENTS 


PART  I 

INTRODUCTION  TO   THE   STUDY   OF   COLLOIDS 

Page 

Introduction xv 

CHAPTER   I 

What  are  Colloids? 3-9 

Sols  —  Suspension,  emulsion,  solution  —  Gels  —  Structure  of  jellies. 

CHAPTER   II 

Surfaces 13-33 

Surfaces  — ■  Chemical  combination  —  Solution  —  Adsorption  —  Sur- 
face pellicles. 

CHAPTER   III 

Size  of  Particles,  Molecular  Weight,  Osmotic  Pressure,  Conduc- 
tivity         41 

CHAPTER  IV 

Phenomena  of  Motion , 49-56 

Brownian  Zsigmondy  movement  —  Diffusion  —  Diffusion  in  jellies 
—  Membranes. 

CHAPTER  V 

Consistency  of  Colloids 64-73 

Internal  friction  —  Swelling  and  shrinking  —  Life  curve  of  colloids. 

CHAPTER  VI 

Optical  and  Electrical  Properties  of  Colloids 75-87 

Optical  properties  —  Electrical  properties  —  Salting  out  —  Floccula- 
tion  — •  Radioactive  substances  as  colloids. 

CHAPTER  VII 

Methods  of  Colloid  Research 89-126 

Dialysis  —  Ultrafiltration  —  Apparatus  —  Pressure  —  Gauging  of 
ultrafilters  —  Adsorption  by  filters  —  Applications  —  Diffusion  in 
aqueous  solution — -Diffusion  in  a  jelly  —  Osmotic  pressure  —  Os- 
motic compensation  method  —  Surface  tension  —  Adsorption  —  In- 
ternal friction  —  Melting,  coagulation  and  solidification  temperatures 

xi 


Xll  TABLE  OF  CONTENTS 

Page 

—  Swelling  —  Flocculation  —  Electric  migration  —  Optical  methods 

—  Interferometer  —  Ultramicroscopy  for  colloidal  solutions  —  Ultra- 
microscopy  for  organized  material. 

PART   II 

THE  BIOCOLLOIDS 
Introduction 129 

CHAPTER  VIII 
Carbohydrates 133 

CHAPTER   IX 
Lipoids 139 

CHAPTER  X 

Proteins 142-166 

Albumins  —  Electrolyte-free  albumin  —  Acid  albumin  —  Alkali 
albumin  —  Albumin  and  inorganic  hydrosols  — -  Albumin,  heavy 
metals  and  salts  of  heavy  metals  —  Globulin  —  Fibrin  —  Nucleins  — 
Albuminoids  — ■  Nucleo  albumins  —  Hemaglobin  —  Colloid  cleavage 
products  of  proteins. 

CHAPTER  XI 

Food  and  Condiments 168-179 

Meat  —  Milk  and  dairy  products  —  Honey  —  Flour,  dough  and 
baking  products  —  Beer. 

CHAPTER  XII 
Enzymes 182 

CHAPTER  XIII 

Immunity  Reactions 193-211 

Nature  of  antigens  and  immune  bodies  —  Distribution  of  immune 
substances  between  suspensions  and  solvent  —  Specific  adsorption  — 
Adsorption  by  organized  suspensions  —  Distribution  of  immune  sub- 
stances between  dissolved  colloids  and  solvents  —  Precipitation  of 
dissolved  colloids  and  organized  suspensions  —  Electric  charge,  H 
and  OH  ions  — ■  Complement  fixation  and  Wassermann  reaction  — 
Wassermann  reaction  —  Anaphylaxis  —  Protective  ferment  —  Meio- 
stagmin  reaction. 

PART   III 
THE  ORGANISM  AS  A   COLLOID   SYSTEM 

Significance  of  the  Colloidal  Condition  for  the  Organism 213 

CHAPTER  XIV 

Metabolism  and  the  Distribution  of  Material 215-245 

Distribution  of  water  in  the  normal  organism  — •  Pathology  of  water 
distribution  —  Edema  —  Inflammation  —  Salt   distribution  —  Cir- 


TABLE  OF  CONTENTS  xiü 

.  •■'         .  Page 

culation  of  material  —  Circulation  of  water  —  Circulation  of  water  in 

animals  —  The  movement  of  water  in  plants  —  Circulation  of  crystal- 
loids —  Circulation  of  coUoids  —  Influence  of  membranes  upon  the 
interchange  of  substances  —  Assimilation  and  dissimilation. 

CHAPTER  XV 

Growth,  Metamorphosis  and  Development 252-274 

Growth  —  Genesis  of  structures — ^  Layered  structures  —  Biological 
growth  —  Ossification  processes  —  Diseases  of  bone  —  Concrements 

—  Gout. 

CHAPTER   XVI 

The  Cell 276-279 

Protoplasm  —  Nucleus  —  Cell  membrane  and  plasma  peUicle. 

CHAPTER   XVII 

The  Movements  of  Organisms 282-292 

Movements  of  lower  organisms  —  Movements  of  higher  organisms 

—  Muscle  as  a  colloid  system  —  Muscle  function  (including  the 
heart). 

CHAPTER  XVIII 

Blood,  Respiration,  Circulation  and  Its  Disturbances 299-315 

Blood  —  Plasma  —  Lymph  —  Blood  corpuscles  —  Respiration  (gas 
exchange)  —  Circulation  and  its  disturbances  —  Secretion  and  ab- 
sorption. 

CHAPTER  XIX 

Absorption 317-323 

Alimentary  absorption  —  Parenteral  absorption. 

CHAPTER  XX 
Secretion  and  Excretion 326-345 

Glands  —  Sahva  —  Bronchial  glands  —  Gastric  juice  —  Secretions 
which  pour  into  the  intestines  —  Ividneys  and  the  secretion  of  urine 

—  Concentration  of  the  glomerular  filtrate  —  Pathology  of  urine 
secretion  —  Result  of  deficient  kidney  function  upon  the  organism  — 
Urine,  normal  —  Urine,  pathological  —  Sweat  glands  —  Milk. 

CHAPTER  XXI 

The  Nerves 352-356 

Nerve  irritability  and  swelhng  —  Cerebrospinal  fluid  —  Integument 

—  Internal  secretions. 

PART   IV 
CHAPTER  XXII 

Toxicology  and  Pharmacology 359-416 

Cooperation  of  indifferent  substances  —  Colloids  —  Adsorption  ther- 
3'Py  — •  Colloidal  metals  —  Action  on  microorganisms  —  Ferments  — 


Xiv  TABLE  OF  CONTENTS 

Page 

Autolysis  —  Metabolism  —  Temperature  curve  —  Distribution  — 
Therapeutics  —  Animal  experiments  —  Clinical  experiments  —  Mer- 
cury —  Sulphur  —  Phosphorus,  arsenic,  antimony  —  Salts  —  Iron 
salts  end  iron  oxid  hydrosol  —  Narcotics  and  anesthetics  —  Disin- 
fection —  Microorganisms  —  Methods  of  testing  disinfectants  — 
Diuretics  and  purgatives  —  Purgatives  —  Astringents  —  Balneology 
—  Water  and  solutions  —  Salves  and  hniments. 

CHAPTER  XXIII 

Microscopical  Technic 417-435 

Maceration  and  isolation  —  Fixing  and  hardening  —  Staining  — 
Theory  of  staining  —  Technic  of  staining  —  The  tissue  elements  in 
their  relation  to  fixatives  and  dyes  —  Staining  of  bacteria. 

Author's  Index 437 

Subject  Index 457 


PART   I. 
INTRODUCTION  TO   THE  STUDY  OF   COLLOIDS. 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


CHAPTER   I. 
WHAT   ARE    COLLOIDS? 

In  spite  of  the  fact  that  they  have  a  much  wider  distribution  than 
crystalloids,  it  is  only  a  little  over  fifty  years  that  colloids  have  been 
scientifically  studied.  Plants  and  animals  and  all  the  things  we  manu- 
facture from  them,  such  as  our  clothing  and  the  greater  part  of  our 
household  goods,  are  colloids.  In  the  year  1861,  Thomas  Graham,*  ^ 
an  Englishman,  called  attention  to  the  fact  that  there  were  substances 
which,  when  in  solution,  diffused  through  parchment  membranes  (dia- 
lysed).  These  he  called  crystalloids  because  the  soluble  crystallizable 
substances  (e.g.,  sugar  and  salt)  possess  this  property  to  a  marked  de- 
gree. Substances  which  were  held  back  by  parchment  membranes 
he  called  colloids,  "glue-like,"  because  glue  was  the  most  character- 
istic of  this  group.  Every  discoverer  of  a  new  fundamental  principle 
is  easily  led  into  exaggerations;  it  so  happened  with  Graham  who 
opposed  crystalloids  to  colloids  as  'Hwo  distinct  worlds  of  matter," 
though  we  know  now  that  all  sorts  of  transition  stages  exist. 

In  succeeding  years  very  few  investigators  concerned  themselves 
with  colloids.  The  very  fruitful  development  of  organic  chemistry 
occupied  the  attention  of  investigators,  who  neglected,  as  less 
important,  a  field  which  promised  fewer  immediate  results.  Only 
in  the  beginning  of  the  new  century  was  there  a  revival  of  interest 
in  colloid  chemistry. 

We  shall  not  follow  the  historic  development  further,  but  shall 
give  a  description  of  colloids  in  accordance  with  the  present  state  of 
the  science.  It  must  be  noted  at  the  outset  that,  even  to-day,  the 
behavior  of  a  dissolved  substance  towards  a  partitioning  membrane, 
that  is,  inability  to  diffuse  through  it,  is  the  chief  characteristic  of 
a  dissolved  colloid. 

Many  colloids  form  with  liquids,  especially  with  water,  a  more  or 
less  fluid  solution.  [The  term  dispersion  is  preferred  by  Arthur  W. 
Thomas  in  a  recent  discussion  on  Nomenclature.  Science,  N.  S., 
Vol.  XLVII,  No.  1201,  p.  10.    Tr.]     This  solution  is  called  a  sol  (from 

1  An  *  after  an  author's  name  refers  to  the  reference  in  the  index  of  authors. 

3 


4  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

solutio  =  solution).  We  speak  of  silver  sols,  albumin  sols,  etc.  The 
dissolved  substance  in  a  sol  may  by  various  means  be  separated  in  an 
amorphous  form  that  retains  more  or  less  water.  This  form  is  called 
a  gel  ^  (from  similarity  to  gelatin) .  If  we  add  salt  to  a  solution  of 
colloidal  silver,  we  obtain  a  black  sediment  containing  very  little 
water,  the  silver  gel.  If  we  boil  a  serum  solution,  the  entire  mass 
solidifies  to  a  jelly  that  does  not  allow  a  separation  of  water  and 
albumin,  the  albumin  gel. 

Sols. 

As  the  researches  of  Graham  have  already  shown,  sols  in  general 
are  substances  which  subdivide  in  their  solvents  into  relatively 
large  particles,  or  which  possess  very  large  molecules,  so  large  that, 
in  contrast  with  the  molecules  of  water  or  crystalloids,  they  are  un- 
able to  pass  through  the  pores  of  an  animal  skin  or  a  parchment 
membrane.  Chemical  grounds  indicate  that  albumin  possesses  a 
very  large  molecule.  Even  though  we  were  to  assume  that  it  split 
into  single  molecules  in  aqueous  solution,  these  are  so  large  that 
they  are  unable  to  pass  through  an  animal  or  vegetable  membrane. 
Accordingly,  the  intact  membranes  of  the  organism  protect  it  from 
loss  of  albumin;  only  in  pathological  conditions  as  in  diseases  of  the 
kidneys  does  albumin  pass  through. 

Substances  like  albumin,^  soluble  starches,  etc.,  are  to  a  certain 
extent  inherently  colloids.  Every  further  subdivision  of  the  colloid- 
ally  dissolved  particles  would  have  to  be  associated  with  a  sphtting 
of  the  molecule,  and  the  fragments  are  certainly  no  longer  albumin, 
but  albumoses,  Polypeptids,  amino-acids,  etc. 

It  is  otherwise  in  the  case  of  certain  artificial  colloids.  Accord- 
ing to  G.  Bredig  and  Th.  Svedberg,  gold,  silver,  platinum  and 
other  metals  may  be  electrically  pulverized  under  water  or  in  organic 
fluids  {e.g.,  isobutyl  alcohol).  According  to  G.  Wegelin,  sihca, 
vanadic  acid,  and  other  substances  may  by  mere  trituration  be 
reduced  to  suspensions  whose  particles  are  so  small  that  they  can- 
not be  recognized  microscopically.  If  the  electrical  pulverization  is 
accomphshed  in  water  which  is  practically  free  from  electrolytes,  we 
obtain  a  solution  which  is  red  in  the  case  of  gold,  brown  in  the  case 
of  silver,  and  greenish  black  with  platinum.     These  solutions  remain 

1  Many  recent  authors  make  "gel"  and  "jelly"  synonymous.  It  seems 
preferable  to  me  to  use  the  expression  "gel"  for  the  general  comprehensive 
phenomenon  and  to  reserve  the  word  "jelly"  for  the  gelatinization  of  a  hydro- 
phile colloid. 

"  When  I  refer  to  albumin,  I  mean  albumin  absolutely  unsplit,  whether  it 
be  egg  albumin  or  globulin,  etc.,  as  opposed  to  albumoses  which  are  classified 
as  albumins  in  some  textbooks. 


WßAT  ARE   COLLOIDS  5 

unchanged  for  months  provided  they  are  preserved  in  Jena  glass, 
which  yields  no  electrolytes  to  water.  These  colloidal  gold,  silver  or 
platinum  solutions  consist  of  more  or  less  fine  metal  particles,  each 
of  which  often  comprises  thousands  of  metal  molecules.  By  varying 
the  strength  and  tension  of  the  current,  finer  or  coarser  particles 
may  be  obtained. 

Gold,  silver  and  other  sols  have  been  prepared  from  gold,  silver 
and  other  salts  by  chemically  liberating  the  metal.  According  to 
the  method  of  preparation,  the  metal  is  obtained  in  a  more  or  less 
fine  state  of  subdivision.  If  the  metal  sol  is  once  produced  it  is 
impossible  by  the  solvent  alone  to  make  the  particles  still  smaller 
(without  using  chemical  means).  Unlike  albumin  they  do  not  have 
the  tendency  to  disintegrate  of  themselves  in  the  solvent.  We  can 
accordingly  call  them  artificial  colloids,  because  they  can  be  brought 
into  such  fine  subdivision  only  by  artificial  means.  If  one  were  to 
further  subdivide  the  molecules  of  such  artificial  colloids,  the  mole- 
cule would  remain  intact;  gold  would  remain  gold,  and  silver,  silver. 
R.  ZsiGMONDY  *  has  prepared  gold  solutions  so  finely  subdivided 
that  they  approach  molecular  dimensions,  and  Th.  Svedberg  has 
shown  that,  with  these  gold  sols,  and  also  with  selenium  sols,  the 
finer  the  subdivision,  the  nearer  these  substances  approach  in  color 
and  light  absorption  their  respective  molecular  solutions.  In  general, 
however,  the  artificial  sols  which  can  be  seen  in  the  ultramicroscope 
consist  of  much  coarser  particles  than  the  natural  sols. 

While  the  chemical  constitution  of  inorganic  colloids  is  revealed 
by  their  method  of  preparation,  nothing  was  known  concerning  the 
constitution  of  natural  organic  colloids  until  1913,  when  Emil  Fischer 
succeeded  in  synthetically  preparing  organic  colloids  resembling 
tannin  and  having  molecular  weights  above  4000.  Exact  knowledge 
of  the  chemical  constitution  of  these  substances  wiU  reveal  much  to 
colloid  research. 

Suspension,  Emulsion,  Solution. 

By  suspension  we  mean  the  floating  of  a  powder  in  a  fluid,  e.g., 
clay  in  water.  An  emulsion  is  the  minute  division  of  one  fluid  in 
another  with  which  it  does  not  mix,  e.g.,  oil  in  milk  or  water.  The 
smaller  the  particles  of  the  "dispersed  phase"  ^  (cf.  p.  11)  of  the  clay 
or  the  fat,  the  longer  it  takes  for  them  to  separate.  Such  a  suspen- 
sion or  emulsion,  in  which  the  dispersed  phase  is  easily  distinguished 

^  Portions  of  a  structure  separated  from  each  other  by  physical  surfaces  are 
called  phases  (Wilh.  Ostwald).  A  mixture  of  oil  and  water  contains  two  phases. 
Oil  is  one  phase  and  water  the  other.  Dispersed  means  scattered,  distributed. 
In  the  above  examples  oil  or  clay  is  the  "dispersed  phase." 


6  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

microscopically,  may  last  months  and  even  years.  Only  about  a 
decade  ago  there  was  an  animated  discussion  as  to  whether  the  known 
inorganic  colloids  such  as  colloidal  silver,  gold,  arsenic  sulphid, 
Prussian  blue,  etc.,  were  suspensions  or  true  homogeneous  solutions. 
Some  evidence  was  against  their  being  considered  homogeneous 
solutions;  other  evidence,  however,  favored  this  view.  Microscopi- 
cally, they  seemed  entirely  homogeneous,  and  they  could  not  be 
separated  from  their  solvents  by  mechanical  means  (filtration  or 
centrif  ugation) . 

It  was  only  by  means  of  the  ultramicroscope,  invented  by  H. 
Siedentopf  and  R.  Zsigmondy  (1903),  that  it  was  convincingly 
shown  that  they  were  suspensions  and  not  homogeneous  solutions. 

After  this  point  was  settled,  the  further  question  arose  as  to 
whether  gelatin,  albumin  sol  and  like  substances  were  to  be  con- 
sidered true  solutions.  Under  the  ultramicroscope,  they  also  showed 
tiny  particles,  which,  however,  were  by  no  means  as  numerous  as 
might  have  been  expected.  Evidently  most  of  them  were  invisible, 
and  it  was  uncertain  whether  this  was  due  to  conditions  of  refrac- 
tion, or  whether  the  larger  part  of  these  substances  was  in  true  solu- 
tion. This  question  was  settled  by  the  method  of  ultrafiltration 
invented  by  H.  Bechhold  in  1906.  By  a  sufficiently  impermeable 
jelly  filter  {ultr  a  filter) ,  that  is,  by  a  purely  mechanical  process,  he  was 
able  to  separate  solutions  of  albumin,  gelatin,  enzymes,  toxins,  etc., 
from  their  aqueous  solvent.  Not  only  did  albumin,  gelatin,  etc., 
prove  to  be  suspensions  or  emulsions,  but  in  addition,  substances 
whose  true  solubility  had  hardly  been  questioned,  e.g.,  most  of  the 
albumoseS;  and  even  dextrin  whose  molecular  weight  had  been  placed 
at  about  1000,  and  which  had  practically  been  classified  as  a  crys- 
talloid. 

It  is  also  possible  to  accomplish  such  a  separation  by  means  of 
centrif  ugation.  By  centrifugation  at  6,000  revolutions  per  minute, 
H.  Bechhold  separated  colloidal  silver  sols  (collargol)  into  coarser 
and  finer  particles.  H.  Friedenthal  has  recently  constructed 
centrifuges  turning  from  10,000  to  30,000  revolutions  per  minute, 
by  means  of  which  he  can  separate  the  casein  from  cows'  milk. 

We  must  here  refer  to  the  definition  for  "homogeneous"  and 
"homogeneous  solution"  given  by  H.  W.  Bakhuis  Roozebom,  whose 
premature  death  we  lament:  "We  call  a  system  homogeneous,  if  all 
its  mechanically  separable  particles  possess  the  sarne  composition  and 
the  same  physical  properties.  Therefore,  this  homogeneity  of  con- 
stitution exists  in  a  well-mixed  liquid  only  because  of  the  smallness 
of  molecules  and  the  coarseness  of  our  means  of  observation." 

We  cannot  speak  of  a  definite  solubility  in  respect  to  suspensions 


WHAT  ARE  COLLOIDS  7 

and  emulsions,  for  within  certain  limits,  we  are  able  to  suspend  as 
much  clay  or  emulsify  as  much  fat  as  we  wish;  the  ''finer"  the  clay 
or  the  fat  is  subdivided,  the  more  "dissolves."  The  same  thing 
holds  for  colloids,  which  are  characteristically  different  in  this  re- 
spect from  crystalloids,  the  latter  having  a  sharply  defined  solubility. 

As  a  matter  of  fact  we  can  get  "supersaturated  solutions"  of 
crystalloids,  and  certain  small  additions  increase  the  solubility  dis- 
proportionately. Such  additions  (e.g.,  albumin,  albumoses,  gelatose, 
dextrin)  when  employed  in  the  case  of  suspensions  and  colloids,  are 
called  protective  colloids  (schutz-kolloide)  because  they  protect  the 
lixiviated  clay  or  finely  dispersed  silver  from  separating  out. 

As  indicated,  many  of  the  pure,  inorganic  sols,  especially  the  metal 
sols  obtained  by  electric  pulverization,  are  very  sensitive  to  elec- 
trolytes by  which  they  are  easily  precipitated,  whereas  on  the  con- 
trary natural  colloids  are  relatively  insensitive.  It  has  been  shown 
that  the  addition  of  certain  natural  sols  acting  as  protective  colloids 
gives  metal  sols,  etc.,  properties  which  cause  them  to  approach  the 
natural  sols  in  stability.  The  inorganic  colloids  employed  in  medi- 
cine, such  as  colloidal  silver  (collargol,  lysargin),  colloidal  calomel 
(kalomelol),  colloidal  bismuth,  etc.,  are  all  stabilized  by  protective 
colloids. 

Thus  we  see  a  complete  transition  from  the  suspension  and  emul- 
sion of  insoluble  substances,  to  the  true  solution  of  crystalloids,  where 
there  occurs  a  disintegration  by  the  solvent,  which  is  so  profound  in 
the  case  of  electrolytes,  that  they  separate  into  their  electrically 
charged  atoms  (ions).  As  everywhere  in  nature,  here  too  there  are 
no  sharp  lines  of  demarcation.  We  cannot  deny  that  at  a  certain 
size  the  particles  possess  the  maximum  colloidal  properties,  especially 
those  conditioned  by  surface  phenomena.  These  properties  decrease 
when  the  particles  are  larger,  i.e.,  if  they  approach  those  of  true  sus- 
pensions or  emulsions;  or  when  they  become  smaller,  i.e.,  if  they 
approach  the  molecular  condition. 

Th.  Svedberg*  has  shown  that  the  light  absorption  of  colloidal 
gold  and  selenium  increases  as  the  particles  become  smaller,  reaches 
a  maximum  in  the  amicroscopic  field,  and  again  decreases  as  the 
particles  approach  molecular  dimensions.  It  is  noteworthy  also 
that  at  a  certain  degree  of  dispersion  the  tinctorial  power  reaches  a 
maximum  which  in  the  case  of  gold  is  forty  times  stronger  than  the 
powerful  color  fuchsin.  The  color  of  colloidal  gold  having  a  particle 
size  of  10  to  20  nn  is  ruby  red;  when  the  particles  are  smaller  it  is 
fuchsin  red;  but  when  the  particles  are  still  smaller  the  color  becomes 
yellowish  red.  In  other  words  it  approaches  the  color  of  gold  salts 
(auric  chlorid)  in  which  the  gold  is  molecularly  dispersed. 


8  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

To  recapitulate:  The  chief  characteristic  of  sols  is  the  large  size 
of  their  particles/  which  are  unable  to  pass  through  vegetable  or 
animal  membranes.  The  natural  size  of  the  particles  accounts  for 
this  in  the  case  of  natural  sols,  while  in  artificial  sols  it  is  due  to  the 
defects  of  our  technic,  which  hitherto  has  not  permitted  our  prep- 
aration of  such  substances  in  molecular,  or  even  approximately- 
molecular  subdivision. 

This  criterion  is  only  valid  for  extreme  cases.  Between  the  un- 
doubted colloids,  e.g.,  albumin,  and  the  undoubted  crystalloids,  e.g., 
amino  acids,  there  are  all  kinds  of  transition  forms,  which  pass 
through  the  same  membranes  more  or  less  rapidly,  e.g.,  albumosea 
and  peptones.  There  is,  indeed,  no  sharp  line  of  demarcation  be- 
tween colloids  and  crystalloids. 

Gels. 

It  might  be  inferred  from  the  nomenclature  (colloids  and  crys- 
talloids) that  the  main  distinguishing  feature  was  the  ability  or 
inability  to  crystallize.  It  is  a  fact  that  most  crystalloids,  i.e.,  sub- 
stances which  pass  through  membranes,  are  cry stalliz able,  whereas 
most  colloids  are  not  able  to  form  crystals  when  they  separate  from 
solution.  However,  this  is  not  a  radical  difference,  since  egg  albumin 
and  hemoglobin  which  are  undoubted  colloids  may  be  obtained  in 
beautiful  crystals;  and  I  have  further  estabUshed  by  ultrafiltration, 
the  colloidal  nature  of  the  solutions  of  the  alkaline  salts  of  the  fatty 
acids  (e.g.,  oleic  acid)  which  also  form  good  crystals.  Colloids  usu- 
ally separate  from  their  solution  in  unformed  masses  called  gels. 

If  the  solid  phase  be  separated  from  crystalloid  solutions,  it  may- 
form  either  crystals  or  a  slightly  or  even  an  entirely  amorphous  pre- 
cipitate. Cuboidal  crystals  separate  from  a  solution  of  common  salt 
on  evaporation  or  addition  of  alcohol,  and  crystals  of  Na2S04  + 
10 II2O  separate  from  solutions  of  Glauber's  salt  (sodium  sul- 
phate). A  white  precipitate  of  barium  sulphate,  which  has  hardly 
any  definite  form,  separates  from  a  sodium  sulphate  solution  upon 
adding  barium  chlorid.  A  substance  of  constant  chemical  com- 
position especially  as  regards  water  content  is  obtained  if  the  im- 
purities, especially  the  extraneous  water,  are  removed  by  filtration 
from  the  crystals  or  precipitate.  To  return  to  our  example:  the 
sodium  chlorid  crystals  and  the  barium  sulphate  are  water-free, 
whereas  the  sodium  sulphate  contains  10  molecules  of  water  to  one 
molecule  of  Na2S04,  but  it  is  water-free  above  33°  C. 

1  By  the  passage  through  a  membrane,  I  mean  especially  passage  by  means 
of  dialysis.  In  many  cases  we  may  substitute  ultrafiltration,  provided  ultrafiltera 
of  sufficient  tightness  are  employed  (see  p.  102). 


WHAT  ARE  COLLOIDS  9 

Gels  behave  differently;  in  fact  there  are  a  number  of  colloids 
which  separate  from  their  solution  almost  water-free.  If  sols  of 
gold,  silver,  platinum,  arsenious  sulphid  or  antimony  sulphid  hydro- 
sol,  prepared  according  to  the  method  of  Bredig  or  the  method  of 
SvEDBERG,  precipitate,  that  is,  separate  from  their  solutions  in  the 
form  of  flocks,  they  are  almost  free  from  water.  Many  inorganic  sols 
{i.e.,  the  artificial  ones),  and  according  to  my  knowledge  nearly  all 
natural  organic  sols,  retain  a  large  quantity  of  water  upon  separation. 

Gelatin  is  the  most  characteristic  gel;  its  aqueous  solutions  (con- 
taining only  1  per  cent  of  water-free  gelatin)  gelatinize  at  ice-box 
temperature.  Furthermore,  other  sols,  such  as  egg  albumin,  starches, 
silicic  acid,  iron  oxid,  etc.,  on  separation  in  gel  form  retain  many 
times  their  own  weight  of  water,  and  form  jelly-like  masses  in  which 
the  proportion  between  the  solid  subsiance  ana  the  retained  solvent 
is  by  no  means  constant.  According  to  the  circumstances  attending 
the  separation,  the  amount  of  water  held  fast  in  the  gel  has  wide 
limits  of  variation.  This  is  a  cardinal  distinction.  In  accord  with 
it,  I  have  adopted  the  happily  chosen  nomenclature  of  J.  Perrin 
and  call  such  colloids  as  throw  down  a  practically  water-free  hydro- 
gel,  hydrophobe  and  those  which  produce  a  hydrogel  swollen  and  rich 
in  water,  hydrophile  colloids.^ 

The  gels  of  hydrosols  (cf.  p.  11)  stabilized  by  protective  colloids 
are  somewhat  hydrophile,  because  very  minute  quantities  of  pro- 
tective colloid  are  sufficient  to  give  the  inorganic  sol  the  properties 
of  the  protective  colloid. 

The  Structure  of  Jellies. 

Jelhes  are  formed  from  their  respective  solutions  by  such  physical 
and  chemical  changes  as  would  cause  the  separation  of  crystals  in 
the  solution  of  a  crystalloid,  e.g.,  by  cooling,  by  removal  of  water 
either  by  a  chemical  change  or  by  forming  an  insoluble  substance  {e.g., 
by  boiling  or  by  acidifying  an  albumin  sol) .  It  is  thus  apparent  that 
jellies  are  to  be  considered  two-phased  structures. 

Two  phases  are  much  more  obvious  in  coagulated  egg  albumen 
whose  opacity  and  white  color  suggest  a  non-homogeneity  of  struc- 
ture. Recently  Bachmann  *  has  demonstrated  ultramicroscopically 
the  two-phased  structure  of  transparent  jellies  such  as  gelatin  and 
silicic  acid.     In  gelatinizing,  it  is  evident  that  granular  flocculent 

1  I  mentioned  above  that  as  far  as  I  knew  all  natural  organic  sols  are  hydro- 
phile. It  might  be  objected  that  epidermis,  hair,  feathers,  bark  and  numerous 
other  vegetable  structures  are  deposited  from  natural  sols  and  become  very 
poor  in  water.  This  is  met  by  the  assertion  that  when  they  are  deposited  they 
contain  much  water  and  that  the  loss  of  water  or  drying  out  occurs  subsequently. 


10  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

and  even  crystalline  particles,  e.g.,  in  soap  jellies,  unite  to  form 
spongelike  structures. 

J.  M.  Van  Bemmelen  compares  the  process  of  separation  of 
colloids  with  the  condition ,  deduced  from  the  phase  rule  govern- 
ing the  separation  of  two  fluids  not  miscible  in  all  proportions,  e.g., 
water  and  phenol.  In  jellies  also,  we  have  a  phase  containing  much 
colloid  and  little  water,  and  another  phase  containing  little  colloid 
and  much  water.  This  conception  of  the  structures  of  jellies  and  the 
process  of  separation  is  due  to  J.  M.  Van  Bemmelen,  O.  Bütschli, 
W.  B.  Hardy,  G.  Quincke,  R.  Zsigmondy  and  W.  Bachmann. 
The  views  of  O.  Bütschli  are  not  accepted  nowadays.  He  main- 
tained that  jelHes  are,  broadly  speaking,  foamy  structures  having 
microscopic  cavities  with  firm  net-like  walls  filled  with  fluid.  Such 
a  structure  can  occur  only  exceptionally. 

The  conception  of  jelhes  as  spongy  structures  gives  us  a  satis- 
factory explanation  of  their  properties.  It  explains  their  solidity 
and  their  plasticity,  their  elasticity,  and  in  short  their  various  physi- 
cal properties. 

The  above  assumption  finds  corroboration  in  another  observation, 
through  the  fact  that  jellies  also  act  as  ultrafilters  and  consequently 
must  be  penetrated  by  fine  capillaries  whose  diameter  has  been 
determined  by  Bechhold  (see  p.  99).  The  penetration  of  jellies 
by  fine  capillaries  filled  with  fluid  was  demonstrated  by  another 
observation.  H.  Bechhold  and  J.  Ziegler*  allowed  salts  which 
would  form  precipitates  with  various  properties  to  diffuse  towards 
each  other  in  gelatin,  e.g.,  potassium  ferrocyanid  and  copper  sulphate 
which  form  a  copper  ferrocyanid  membrane  entirely  impervious  to 
electrolytes;  silver  nitrate  and  sodium  chlorid  which  form  a  silver 
chlorid  membrane  which  is  permeable  for  electrolytes  if  the  osmotic 
pressure  is  higher  on  one  side  than  on  the  other.  Microscopic  sections 
through  the  membranes  formed  by  the  precipitates  prove  that  the 
gelatin  is  not  deformed.  Accordingly,  when  diffusion  ceases,  it  is 
because  the  diffusion  paths  have  been  obstructed,  i.e.,  a  precipitate 
has  been  formed  in  the  fluid  phase  so  that  the  paths  are  closed  and 
of  course  the  gelatin  walls  which  contain  little  water  are  impassable 
for  electrolytes.  Remelting  is  sufficient  to  reopen  the  diffusion 
paths.  Anderson  determined  a  diameter  of  5.2  fxfj.  for  the  largest 
pores  of  silicic  acid  jelly  from  the  vapor  pressure  reduction  which  a 
fluid  undergoes  in  cylindrical  capillaries. 

Until  now  we  have  assumed  that  sols  exist  only  a§  aqueous  solu- 
tions and  that  there  are  only  aqueous  gels.  This  is  by  no  means 
true.  Even  Thomas  Graham  *  showed  that  water  could  be  replaced 
by  alcohol  and  glycerin.     Th.  Svedberg  pulverized  numerous  metals 


WHAT  ARE  COLLOIDS  11 

in  organic  fluids  especially  in  isobutyl  alcohol.  R.  Lorenz  has  even 
made  metal  sols  in  red  hot  solution  (pyrosols)  by  electrolysis  of 
molten  lead  and  cadmium  salts,  etc.  To  distinguish  them  from  the 
water  soluble  hydrosols  and  from  the  hydrogels  we  call  them  organo- 
sols or  organogels  and  according  to  the  solvent  as  alcosols,  etc.  These 
do  not  occur  in  nature  and  are  therefore  of  no  importance  to  us. 

An  investigation  of  such  colloids  as  have  fats,  lecithin  and  Cholesterin 
either  as  a  dispersing  medium  or  as  a  "dispersed  phase"  (cf.  p.  12) 
would  certainly  be  of  great  importance  to  biology  and  medicine. 
The  fact  that  fats  and  oils,  especially  mineral  oils,  may  serve  as  a 
dispersing  medium  for  colloids  has  been  mentioned  by  D.  Holde  * 
in  his  work  on  the  "physical  condition  of  solid  fats"  and  this  was 
confirmed  by  investigations  of  H.  Bechhold,i  who  was  able  upon 
ultrafiltration  (through  a  toluol-glacial  acetic  acid  collodion  filter) 
to  hold  back  from  a  crude  oil  a  part  of  the  asphalt  colloidally  dis- 
solved in  it.  Recently  C.  Amberger  dissolved  a  series  of  metals 
(gold,  silver,  platinum,  arsenic,  etc.)  in  lanolin.  Some  of  these 
solutions  have  therapeutic  application. 

By  ultrafiltration  H.  Bechhold  separated  from  commercial 
chlorophyl  the  coloring  matter  and  the  wax-like  products  which  were 
evidently  held  in  colloidal  solution. 

Protective  colloids  also  exist  for  organic  fluids.  Iron  oxid  gel  and 
iron  oxid  hydrosol,  rennin  and  trypsin,  as  well  as  albumoses  which 
are  completely  insoluble  in  chloroform,  become  soluble  in  it  with  the 
aid  of  lecithin  acting  as  a  protective  colloid. 

Thus  far  we  have  sought  to  obtain  a  picture  of  what  we  term 
^'colloids"  and  now  we  shall  strive  to  elucidate  upon  what  their 
properties  depend.  In  solutions  of  cofloids  and  in  gels,  we  have 
mixtures  of  solids  or  of  fluids  with  fluids.  It  has  for  a  long  time 
been  known,  that  at  the  interface  between  two  substances  which  do 
not  mix  (air  and  water,  oil  and  water,  glass  and  water)  there  occur 
phenomena,  called  surface  phenomena.  For  instance,  the  surface  of 
water  in  contact  with  air  acts  as  a  pellicle;  if  we  allow  water  to  drip, 
each  drop  reaches  a  considerable  size  before  its  weight  breaks  through 
the  surface  skin  or  pellicle  and  the  drop  falls.  This  surface  skin  is 
much  weaker  in  the  case  of  alcohol,  so  that  drops  of  alcohol  falling 
from  the  same  tube  are  much  smaller  than  those  of  water.  The  fol- 
lowing is  another  example  of  a  surface  phenomenon:  oil  forms  a 
sphere  in  a  suitable  mixture  of  water  and  alcohol;  if  we  raise  the 
specific  gravity  of  the  water  by  removing  some  of  the  alcohol,  the 
oil  rises  and  spreads  out  over  the  surface  of  the  water. 

Such  surface  phenomena  are  very  numerous;  they  are  brought 
1  Unpublished. 


12  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

about  by  the  fact  that  different  conditions  exist  in  the  interior  than 
exist  on  the  surface  of  the  fluid  or  sohd  body.  In  two-phase 
systems,  such  as  colloids,  in  which  the  interfaces  reach  enormous 
dimensions,  surface  and  capillary  phenomena  become  most  promi- 
nent; in  fact,  they  are  in  many  ways  the  most  characteristic  phe- 
nomena of  colloids.  Two  fluids  which  do  not  mix  are  described  as 
two  fluid  ''phases";  it  is  possible  also  to  speak  of  a  fluid,  of  a  solid 
and  of  a  gaseous  phase.  In  order  to  give  expression  to  the  great 
surface  development  in  a  sol  or  gel,  Wolfgang  Ostwald  introduced 
the  very  happy  expression  dispersed  phase.  In  a  silver  sol,  silver 
is  the  soUd  dispersed  phase;  in  an  oil  emulsion,  oil  is  the  fluid  dis- 
persed phase;  in  both,  water  is  the  dispersing  medium.  Colloidal 
solutions  and  gels  are  all  dispersed  systems. 

[Of  interest  in  this  connection  is  the  work  of  G.  H.  A.  Clowes  and 
Martin  H.  Fischer.     Tr.] 


CHAPTER  II. 


SURFACES. 

We  have  seen  in  the  preceding  chapter  that  both  colloidal  solutions 
and  jellies  are  to  be  regarded  as  two-phase  systems.  In  dispersed 
systems,  the  surfaces  of  contact  acquire  an  overpowering  importance 
by  reason  of  their  enormous  development. 

In  order  to  get  an  idea  of  the  increased  development  of  surface 
attending  progressive  subdivision,  I  give  below  a  table  taken  from 
Wolfgang  Ostwald: 


Edge  length 

Number  of  cubes 
occupying  a  vol- 
ume of  1  cm.3 

Total  surface 

1          cm 

0.1      cm 

0.01    cm 

1 

10^ 
10« 

10« 

1012 
1015 
1018 

1021 
1024 

6  cm. 2 

60  cm. 2 

600  cm. 2 

0.001  cm. 

(The  diameter  of  a  human  blood  corpuscle 
is  about  0.0007  cm.) 

6,000  cm. 2 

Im 
0.1m 

(  =  0.0001  cm.;  diameter  of  a  small  coccus). 

6m.2 
60m.2 

0.01m 
1mm 

(Limit  of  ultramicroscopic  visibility) 

600  m.2 

(The  diameter  of  the  finest  colloidal  parti- 

6,000  m.2 
60,000  m.2 

0.1mm 

(Diameter  of  elemental  molecules) 

From  this,  we  can  understand  how  small  surface  forces  may,  in  a 
dispersed  system,  become  most  important,  and  mask  other  phe- 
nomena. We  shall  proceed  to  the  study  of  the  properties  of  sur- 
faces in  the  following  pages. 

If  we  compare  a  point  A  (Fig.  1)  in  the  interior  of  a  phase,  e.g., 
a  fluid,  with  one  on  the  surface  A',  e.g.,  in  contact  with  air,  we  notice 
that  the  former  is  surrounded  on  all  sides  by  an  impervious  mass, 
whereas  the  latter,  surrounded  on  only  one  side  by  such  a  mass, 
experiences  an  attraction  to  the  fluid  phase.  The  pressure  with 
which  the  surface  layer  is  drawn  inwards  is  called  inward  attraction 
(Jbinnendruck) . 

If  we  imagine  a  drop  of  water  to  be  on  a  surface  which  it  does  not 
moisten,  e.g.,  a  leaf,  there  is  a  pulling  towards  the  center  from  all 
sides,  which  means  that  the  drop  takes  a  spherical  form,  in  order 
that  it  may  have  the  smallest  possible  surface.     The  surface  acts 

13 


14 


COLLOIDS  IN  BIOLOGY  AND   MEDICINE 


like  an  elastic  skin  which  surrounds  the  drop.  If  we  drop  water 
from  a  tube,  the  drop  develops  in  size  and  is  retained  by  its  skin 
until  the  increasing  weight  tears  it  away.^  We  call  this  force  which 
determines  the  tension  of  the  surface,  the  surface  tension  (a).  A  flat 
surface  of  water  of  1  square  cm.  endeavors  to  contract  with  a  tr  of 
about  0.075  gm.,  when  it  is  spread  out  like  a  soap  bubble  with  air 


Air  interface 


Fluid 


Fig.  1. 


pressure.  Every  change  in  shape  of  the  sphere  of  water,  i.e.,  every 
increase  of  the  surface,  presupposes  work;  this  depends  upon  the 
surface  area  (co)  and  the  surface  tension  (a).     Surface  energy  =  a '  (ji 


surface  energy 


dynes 


or  (7  = ^^  expressed  m 

CO  cm. 

There  are  many  methods  for  determining  the  surface  tension. 
Some  of  them  depend  upon  the  shape  (distortion)  which  the  sur- 
face of  a  fluid  takes;  some,  upon  the  height  to  which  a  fluid 
ascends  a  capillary;  some,  upon  the  determination  of  the  maximum 
weight  attained  by  a  falling  drop.  Detailed  descriptions  are  to  be 
found  in  every  large  treatise  on  Physics,  such  as  H.  Freundlich's 
"  Kapillarchemie." 

A  number  of  values  of  a  are  given  as  examples.  The  a  of  water  is 
determined  most  frequently;  it  has  the  highest  a  of  all  the  sub- 
stances which  are  fluid  at  room  temperature  (mercury  excepted). 
The  various  methods  give  quite  divergent  values.  The  values  given 
here  are  in  each  case  the  surface  tension  towards  air. 


(fluid/air) 
<r 

Water 71.7-76.8 

Mercury 436 

Benzol 28.8 

Ethyl  alcohol 22 

Ethyl  ether 16.5 

Glycerin' 65 

Acetic  acid 23 . 5 

^  The  comparison  to  an  elastic  membrane,  at  best,  has  only  a  limited  applica- 
biUty,  since  it  is  true  that  the  surfaces  are  enlarged  or  diminished,  yet  they  are 
not  stretched.  The  particles  in  the  surface  neither  separate  nor  approach  each 
other  but  more  particles  are  forced  into  the  surface  (when  the  surface  enlarges) 
or  removed  from  it  when  it  diminishes. 


(fluid /air) 
a 

n-Butyric  acid 26. 3 

Chloroform 26 

Ohve  oil 32.7 

Resin  (melted  and  sohdified)     37.2-33.1 

Glue 48.3 

SheUac 36.7-30.4 


SURFACES  15 

The  gas  particles  on  the  surface  between  fluid/air  are  affected  by 
different  forces  than  in  the  center  of  the  gas  space;  they  receive  a  pull 
in  the  direction  of  the  fluid  phase,  they  are  condensed  at  the  inter- 
face, and  in  the  surface  film,  fluid  is  constantly  changing  into  gas. 
Analogous  phenomena  occur  wherever  interfaces  are  formed,  that  is, 
at  the  interface  between  fluid/gas,  fluid/fluid  (in  the  case  of  non- 
miscible  fluids),  gas/solid,  fluid/soKd,  solid/solid.  At  the  fluid/gas 
and  fluid/fluid  interfaces  we  recognize  these  forces  by  the  shape 
of  the  surface  of  contact  (concave  or  convex),  which  is  formed 
under  the  influence  of  the  surface  tension.  At  the  interface 
between  solid/gas  we  recognize  the  condensation  of  the  gaseous 
phase  at  the  surface  by  the  fact  that  it  is  almost  impossible  to 
remove  the  layer  of  gas#^  This  is  the  reason  why  it  is  so  difficult 
to  create  high  vacua,  and  why  exhaustion  of  gas  is  undertaken  in 
the  presence  of  charcoal  which  by  reason  of  its  still  greater  surface 
removes  that  layer  of  gas  which  clings  to  the  glass  vessel.  Spongy 
platinum  acts  as  an  igniter  by  condensing  on  its  surface  gases  which 
then  unite  chemically  with  the  liberation  of  heat. 

The  surface  tension  at  the  interfaces  between  two  fluids  can  be 
determined  by  the  same  methods  that  are  employed  in  the  case  of 
fluid/gas.  In  this  case  also,  a  limiting  surface  develops  at  the 
interface  of  the  two  phases,  which  changes  with  every  variation  of 
surface  tension. 

A  few  values  of  a  for  fluid /fluid  are  now  given: 

<r 

Water/benzol 32. 6 

Water/oil  of  turpentine 12. 4 

Water/chloroform 27 . 7 

Water /ethyl  ether 9. 69 

Water /isobutyl  alcohol 1 .  76 

Water /rosin 19.8 

Water /olive  oil 22 . 9 

Alcohol/olive  oil 2 .  26 

Rape  seed  oil/egg  albumen 7. 10 

OHve  oil/ox  gall  (9  per  cent) 7. 21 

Ohve  oil/Castile  soap  (1/4000) 3 .  65 

Olive  oil/rubber  solution 14 . 9-10 . 2 

A  method  for  measuring  the  surface  tension  of  the  dispersed  phase 
in  an  emulsion  is  here  described  in  some  detail,  because  it  is  too 
new  to  be  found  as  yet  in  any  textbook  on  Physics,  and  particularly 
because  on  account  of  its  general  applicability  it  promises  to  be  of 
especially  great  importance  for  colloid  chemical  research. 

E.  Hatschek  *  separated  an  oil  emulsion  into  water  and  oil 
by  means  of  idtrafiltration.  From  these  experiments  Hatschek 
deduced  the  diameter  of  drops  of  oil  and  the  size  of  pores  of  the 


16 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


ultrafilter.  The  author  raised  certain  objections  to  these  conclusions, 
as  it  was  not  necessary  to  assume  that  the  drops  of  oil  retained  their 
form  in  passing  through  the  filter  pores;  they  might  have  been  drawn 
out  into  thread-hke  cyhnders,  and  their  diameter  thereby  reduced. 
As  a  result  of  the  correspondence  that  followed,  H.  Bechhold  pro- 
posed that  the  pressure  necessary  to  change  the  shape  of  a  sphere  of 
oil  in  a  fluid  be  estimated  and  experimentally  verified. 

E.  Hatschek*  carried  out  these  calculations  and  measurements. 
The  method  of  calculation  is  in  brief  the  following :  Upon  entering  a 
capillary,  a  sphere  changes  into  a  cylinder  which  is  bounded  above 
and  below  by  a  meniscus;  to  simplify  matters  these  meniscuses  are 
regarded  as  hemispheres.     Accordingly,  let 

R  =  radius  of  the  oil  sphere, 
r  =  radius  of  the  capillary, 
then  R  =  nr; 

a  =  surface  tension  in  dynes/cm., 
p  =  pressure  per  surface  unit  of  the  emulsion, 
g  =  980  (acceleration  due  to  gravity), 
an 


then, 


P 


Rg 


n  approximately    =  C  (n  —  1),   so  that  C  lies  between  1.8  and 

aC  {n  -  1) 


Rg 
pRg 
Ca 
pRg 


;  or,  if  we  desire  to  de- 
+  1 ;  or,  should  we  desire  to 


1.9,  and  we  have  the  equation  p  = 
termine  the  size  of  the  pores,  n  = 

determine  the  surface  tension  a-  =  „  ,         . . 

(7  (n  —  1) 

E.  Hatschek  tested  the  correctness  of  the  formula  by  determining 
the  pressure  necessary  to  deform  a  sphere  of  mercury  or  oil  (nitroben- 
zol)  sufficiently  to  make  it  enter  a  narrow  capillary  and  obtained 
satisfactory  results.  To  give  an  idea  of  the  pressures  that  are  in- 
volved, let  us  consider  the  following  example:  In  order  to  force  a 
drop  of  mercury  with  a  radius  of  0.111  cm.  in  water  into  a  capillary 
with  a  radius  r,  a  water  colunm  of  p  centimeters  was  necessary. 


r  (in  cm.) 

p  per  cm.  of  vessel 
(calculated) 

p  (observed) 

0.0255 
0.0112 

21.9 
58.65 

20.5 
62.0-63.2 

In  an  emulsion  of  oil  in  water  in  which  the  diameter  of  the  oil  drops 
was  0.4  fjL,  a  pressure  of  20  atmospheres  was  required  to  press  them 
through  pores  having  a  diameter  of  75  mm-     According  to  this  theory 


SURFACES  17 

it  should  be  possible  to  obtain  a  clear  or  a  cloudy  filtrate  from  an  oil 
emulsion  according  to  the  pressure  employed.  This  assumption  was 
confirmed  by  an  experiment  of  H.  Bechhold:  When  an  emulsion  of 
oil  in  water  was  filtered  through  a  3  per  cent  ultrafilter  with  a  pres- 
sure of  6  atmospheres  a  clear  filtrate  was  obtained,  and  when  the 
pressure  was  increased  to  10  atmospheres  the  filtrate  became 
turbid;  with  a  decrease  in  the  pressure  the  filtrate  became  clear 
again. 

In  my  opinion  the  greatest  importance  of  the  method  lies  in  the 
ability  to  measure  the  surface  tension  of  small  fluid  or  semifluid 
structures  (e.g.,  blood  corpuscles)  and  to  deduce  from  such  determi- 
nations entirely  new  points  of  view  concerning  the  passage  of  fluid 
or  semifluid  structures  through  membranes. 

Let  me  point  out  another  idea  which  forces  itself  upon  me :  namely, 
that  the  sphere  is  the  form  most  readily  induced  by  surface  tension. 
If  a  solid  substance  separates  from  a  fluid  as  a  crystal  we  must 
recognize  that  certain  forces  tending  to  increase  the  surface  are 
opposing  the  surface  tension.  But  we  know  from  the  microscopic 
study  of  crystal  formation  that  spherical  structures  usually  appear 
first;  later,  crystalline  forms  with  rounded  corners,  and  only  in  the 
later  stages  true  crystals.^  There  must  be  a  certain  relation  between 
mass  and  surface  in  order  that  the  solid  phase  may  be  elevated 
above  the  surfaces  bounded  by  planes  in  defiance  of  the  surface 
tension.  If  the  surface  is  too  great  in  proportion  to  the  mass,  the 
surface  tension  overcomes  the  crystallizing  forces.  Since  it  is  possi- 
ble to  estimate  the  increase  in  surface  acquired  by  the  identical 
substance  in  changing  from  a  spherical  form  to  a  crystal,  and  further, 
to  observe  the  smallest  quantity  of  a  substance  which  can  become 
crystalline,  it  becomes  possible  to  solve  many  problems,  such  as,  the 
effective  forces  of  crystallization,  the  surface  tension  which  solid  bodies 
exert  against  their  solutions,  and  the  decreased  capacity  to  crystal- 
lize in  the  presence  of  colloids. 

A  drop  of  oil  spreads  out  over  the  surface  of  water.  This  occurs  be- 
cause the  surface  tension  of  water  acting  against  oil  is  less  than  the 
surface  tension  of  the  water  acting  against  air  minus  the  surface 
tension  of  the  oil  acting  against  air.  (The  explanation  of  this  is 
given  in  all  the  larger  text  books  on  Physics.) 


water/air 

> 

(T  water/oil 

+ 

a  oil/air 

75 

> 

22.9 

+ 

32.7. 

1  Bibliography  in  Wo.  Ostwald,  "  Handbook  of  Colloid  Chemistry,"  trans,  by 
Fischer,  Oesper  and  Berman,  Phila.,  1915. 


18  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Expressed  in  general  terms,  this  means  that  a  fluid  2  spreads  itself 
on  the  free  surface  of  a  fluid  1,  if 

O"!  >0'2  +  0'l/2- 

(Ti  =  surface  tension  of  fluid  1  acting  against  air. 
0-2  =  surface  tension  of  fluid  2  acting  against  air. 
(Ti/2  =  surface  tension  of  fluid  1  acting  against  fluid  2. 

Similarly,  a  fluid  3  spreads  over  the  common  boundaries  of  two 
fluids  1  and  2,  whenever 

0'l/2  >   ö'2/3  —  (Ts/l. 

This  phenomenon  is  of  the  greatest  biological  interest,  because  it 
follows  that  many  fluids  must  spread  out  on  the  boundaries  of  other 
fluids  or  solid  bodies  and  form  films,  and  further,  that  solid  particles, 
suspensions  and  colloids  must  collect  on  surfaces,  according  to  con- 
ditions. The  diminution  of  surface  tension  between  two  surfaces  is 
the  forerunner  of  mixing  or  solution;  there  exists  no  surface  tension 
between  two  readily  miscible  fluids.  We  shall  discuss  the  distribu- 
tion of  dissolved  substances  on  interfaces  more  fully  on  page  33  in 
the  discussion  of  surface  pellicles.  We  shall  see  that  numerous 
organized  structures  and  indeed  the  movement  of  protoplasm  and 
of  the  lower  animals  are  derived  from  this  phenomenon  of  surface 
tension.  It  is  the  monumental  service  of  G.  Quincke  that  he  showed 
the  connection  between  the  purely  physical  processes  and  the  phe- 
nomena of  animate  nature. 

The  surface  tension  of  solids  is  deduced  from  the  fact  that  finely 
divided  particles  are  more  easily  and  rapidly  dissolved  than  coarser 
ones;  it  also  explains  the  fact  that  artificial  hydrophobe  colloids  are 
produced  only  in  a  dispersing  medium  in  which  they  are  wholly  in- 
soluble (see  p.  73).  The  slightest  solubility  permits  them  to  pass 
from  the  dispersed  phase  into  coarser  particles.  Only  to  a  limited 
extent  is  it  possible  directly  to  measure  the  surface  tension  of  solid 
bodies.  Roentgen  measured  the  a  for  rubber/air  and  rubber/water 
and  Tangl  *  tested  a  new  method  on  the  interfaces  of  rubber/water 
and  paraffin/ water.  The  basis  for  the  method  is  that  a  tube  of  the 
substance  to  be  tested  (rubber  or  paraffin)  undergoes  a  change  in 
shape  when  it  is  plunged  from  the  air  into  a  fluid  (water) . 

Interfaces  of  Solutions.  Whenever  substances  are  dissolved  in 
one  or  both  phases,  there  is  usually  a  difference  between  the  con- 
centration on  the  surface  and  on  the  interior.  These  changes  in 
concentration  at  the  surface  are  termed  adsorption.  A  substance  be- 
comes concentrated  upon  the  surface  if  it  reduces  the  surface  tension. 
This  is  the  most  usual  adsorption  phenomenon.     Only  a  few  inor- 


^       SURFACES  19 

ganic  salts  increase  the  surface  tension  of  water  and  they  are,  ac- 
cordingly, less  concentrated  at  the  surface  than  in  the  interior  of 
the  solution.  This  latter  occurrence,  negative  adsorption,  is  of 
significance  for  the  distribution  of  salts  in  cells,  as  will  be  shown  on 
page  25. 

''Adsorption,"  which  is  of  the  greatest  significance  in  colloid 
research,  will,  in  the  subsequent  paragraphs,  be  considered  from  the 
standpoint  of  the  distribution  of  a  dissolved  substance  between  two 
phases.  In  this  connection  also  reference  should  be  made  to  page 
33  (surface  pellicles). 

CHEMICAL   COMBINATION,   SOLUTION,  ADSORPTION. 

We  have  seen  that  colloids  are  diphasic  systems,  and  the  ques- 
tion must  arise  as  to  how  a  third  substance  will  he  distributed  between 
the  two  phases.^ 

Chemical  Combination. 

If  we  take  as  the  dispersed  phase,  a  suspension  of  calcium  carbon- 
ate (precipitated  chalk)  in  water,  this  will  represent  the  solution  of  a 
hydrophobe  colloid.  If  we  add  sulphuric  acid  to  the  suspension,  the 
acid  will  be  immediately  and  completely  bound.  It  is  impossible  to 
detect  free  sulphuric  acid  in  the  suspension  by  any  reagent  if  any 
calcium  carbonate  still  remains  in  the  suspension.^  If  we  continue 
with  the  addition  of  sulphuric  acid,  a  point  will  suddenly  occur  when, 
no  matter  how  much  is  added,  sulphuric  acid  is  no  longer  bound  by 
the  chalk;  all  the  excess  remains  in  the  water.  We  are  accustomed 
to  say  that  a  chemical  reaction  has  occurred  between  the  calcium 
carbonate  and  the  sulphuric  acid  and  that  there  is  a  chemical  union 
of  Ca  and  SO4.  Ca  unites  firmly  with  a  definite  quantity  of  SO4. 
We  may  add  as  much  water  as  we  want;  it  cannot  abstract  any 
S04from  the  CaS04.     The  process  is  irreversible  (cannot  be  reversed). 

Solution. 

If  we  emulsify  carbon  disulphid  in  water,  and  add  a  little  bromin, 
the  entire  fluid  is  colored  brown.  If  we  allow  the  carbon  disulphid 
to  settle,  the  water  is  light  brown  and  the  carbon  disulphid  is  colored 

^  In  this  instance  "distribution"  is  used  in  the  most  general  sense,  though  in 
physical  chemistry  it  is  employed  only  to  indicate  the  distribution  of  a  sub- 
stance between  two  solvents. 

2  Although  the  formation  of  salts  on  mixing  dissolved  acids  and  bases  results 
with  infinite  rapidity,  it  takes  an  appreciable  time  in  the  case  of  colloid  solutions 
(and  of  course  with  coarse  suspensions).     Vorländer  and  Häberle, 


20  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

dark  brown.  The  more  bromin  we  add,  the  darker  both  the  water 
and  the  carbon  disulphid  become;  the  latter,  however,  is  always 
more  darkly  colored  than  the  former.  If  we  study  the  process 
quantitatively,  the  following  becomes  evident:  if  in  a  given  case  the 
concentration  of   the   bromin  in  the  carbon   disulphid  is  c  (carbon 

Til.,..,,           ^         /      j^    N     ^1         cfcarbon  disulphid) 
disulphid),   m  the   water  c (water),   then  -, — - — r-^ =  n, 

that  is,  the  relative  distribution  of  the  bromin  in  a  given  case  is  n. 

If  we  double  the  quantity  of  both  carbon  disulphid  and  of  water 
and  then  test  the  quantity  of  bromin  in  the  fluids,  we  shall  find  that 
in  both  the  concentration  has  fallen  to  half  and  that  the  distribution 
continues  to  be  n.  If  we  double  the  quantity  of  water,  its  color  is 
only  slightly  less  intense,  because  bromin  enters  the  water  from  the 
carbon  disulphid.  If  we  now  measure  the  bromin  content  of  the  two 
fluids,  we  shall  find  ac (carbon  disulphid)  and  in  the  water  &c (water); 
that  is,  ac  (carbon  disulphid)  _ 

6c  (water) 

No  matter  how  we  vary  the  quantity  of  solvent  or  of  bromin,  the 
apportionment  of  bromin  is  always  n.     We  may,  therefore,  say  that 

c 

n  is  a  constant  and  express  it  —  =  k. 

Ci 

This  equation  is  characteristic  for  the  distribution  of  a  substance 
between  two  phases  in  which  it  is  soluble.  The  process  is  reversible; 
it  is  in  labile  balance.  The  law  of  distribution  was  formulated  by 
M.  Berthelot  and  Jungfleisch  in  1872,  though  we  still  frequently 
refer  to  Henry's  Law  of  Distribution.  Strictly  speaking,  this  ex- 
pression applies  only  to  the  distribution  of  a  gas  between  a  fluid 
and  a  gaseous  phase  (proportionately  to  the  pressure). 

c  .  . 

The  distribution  —  =  k  applies  only  in  case  the  molecular  weight 

of  the  dissolved  substance  is  the  same  in  both  solvents.     If  this  is 

c 

not  the  case,  the  equation  becomes  —  =  k,  in  which  a  and  b  express 

the  difference  in  the  molecular  weight  in  the  two  solvents.^  W. 
Nernst  has  formulated  the  law  of  distribution  in  this  way.  For 
example,  benzoic  acid  in  water  has  a  simple  molecular  weight,  but 
in  benzol  it  exists  mostly  in  double  molecules.  In  order  to  ex- 
press this,  the  equation  of  distribution  becomes     .  =-  =  k. 

VC  (benzol) 

1  [Prof.  J.  L.  R.  Morgan  suggests  that  a  better  form  would  be  the  following: 

—  =  k, 

where  x  is  the  ratio  of  the  molecular  weight  in  solvent  (2)  to  that  in  solvent  (1). 

Sx  ^  xs      and       — -. — TTT-  =  X.    Tr.j 
(2)     (1)  m  m  (1) 


SURFACES  21 

Adsorption. 

With  colloids  in  particular  there  occurs  a  third  possibiUty  of  dis- 
tribution, wherein  the  surface  comes  into  play  rather  than  the  total 
mass  of  the  dispersed  phase.  The  condition  of  distribution  which 
we  are  about  to  describe  is  called  adsorption.  Suppose  we  suspend 
in  water  a  substance  which  we  may  assume  does  not  dissolve  or 
undergo  chemical  combination,  e.g.,  pure  carbon.  We  know  that 
bone  black  may  to  a  greater  or  less  extent  decolorize  dye  solutions; 
that  it  is  used  to  bleach  dark  sugar  juices  and  to  decolorize  the  dark 
solutions  of  the  organic  chemist.  When  suspension  of  powdered  char- 
coal is  added  to  bromin  water,  we  observe  the  following:  If  we  add 
very  little  bromin  to  the  water  the  latter  will  become  completely 
decolorized;  if  we  add  more,  a  considerable  part  is  taken  up  by  the 
charcoal  but  the  water  becomes  brownish.  With  further  addition 
of  bromin  the  water  is  colored  more  intensely  and  the  charcoal  takes 
up  proportionately  less  bromin.  This  process  is  reversible  and  the 
distribution  of  bromin  between  charcoal  and  water  follows  a  cer- 
tain law.  We  cannot  as  in  the  case  of  a  solvent,  however,  speak 
of  the  concentration  of  the  dispersed  phase.  In  experiments,  it  has 
become  customary  to  insert  the  specific  gravity  of  the  dispersed 
phase,  and  this  custom  is,  as  a  rule,  justified.  Let  us,  for  example, 
designate  by  x  the  amount  in  millimols  of  bromin  that  is  adsorbed 
from  a  solution  by  m  grams  of  charcoal,  and  by  c,  the  concentration 
of  the  bromin  in  the  water  after  adsorption.  If  we  deal  with  sub- 
stances of  unknown  molecular  weight,  x  indicates  the  weight  in  milli- 
grams and  c  the  weight  which  is  present  in  1  cc.  of  water  after 
determining  the  adsorption  balance.  Empirically  we  arrive  at  the 
equation 

—  (adsorbed) 
m 

T =  /«, 

c"  (free) 

in  which  the  exponent  -  is  always  <  1.     Inspection  shows  that  if 

X 

n  =  3  and  k  =  20,  the  equation  is  satisfied  when  —   in  the  char- 

m 

coal  =  200  and  c  (water)  =  1000. 

If  we  dissolve  but  very  little  bromin,  then  the  equation  is  satisfied 

X 

when,  e.g.,—  =  20  and  c  =  1.     In  the  first  instance  only  1/5  of  the 

bromin  is  adsorbed  by  the  charcoal,  but  in  very  great  dilutions  20 
times  as  much  goes  to  the  adsorbent. 


22 


COLLOIDS   IN  BIOLOGY  AND   MEDICINE 


If  it  is  unnecessary  to  determine  constants,  the  direct  graphic 
representation  of  results  is  the  simplest  method.  The  concentra- 
tion in  water  (the  dispersing  substance)  is  made  the  abscissa.  The 
ordinate  is  the  adsorbed  amount  of  the  dispersed  phase.  (This  is 
the  difference  between  the  entire  quantity  of  the  substance  dis- 
solved and  what  remains  in  solution.)  The  points  of  intersection 
are  points  of  the  curve  experimentally  derived.  The  lines  show  us 
at  a  glance  (as  is  seen  in  Fig.  2),  in  simple  cases,  whether  the  dis- 


cs 
a, 


^' 

^' 

-  / 

f 

/ 
/ 

,^ 

/ 

/ 
/ 

/ 

/ 

/ 

1 

i 
i 

/ 

/ 

/ 

/ 
/ 

j        / 

/ 
1   / 
I  / 

C  (concentration)  in  the  water 
Fig.  2. 

tribution  and  curve  is  one  of  chemical  combination,  solution  or 
adsorption. 

The  heavy  continuous  line  ( )  is  the  graphic  representation  of 

a  chemical  reaction:  3  mols  CaCOs  are  suspended  in  water  and 
H2SO4  is  added.  It  is  at  once  seen  from  the  diagram  that  3  mols 
II2SO4  are  bound,  i.e.,  there  is  no  free  H2SO4  in  the  water,  but  on  the 
addition  of  more  II2SO4  the  dispersed  phase  can  take  up  no  more 
acid,  so  that  the  acid  remains  in  the  dispersing  medium. 

The  broken  line  ( )  is  the  graphic  representation  of  the  dis- 
tribution of  a  substance  between  two  solvents.  The  dot  and  dash 
line  (-  •  -)  is  an  adsorption  curve.  For  the  graphic  representation  of 
such  adsorption  phenomena  the  above  equation  is  solved  by  log- 
arithms;  and  we  obtain 

X  1 

log  —  =  log  fc  4-  -  •  log  c. 

m  n 


SURFACES 


23 


This  is  the  equation  of  a  Hne.     If  the  logarithms  of  the  values  found 

for  —  and  for  c,  of  different  concentrations  of  the  substances  under 
,    ^  m 

examination,  are  carried  onto  a  rectangular  system  of  co-ordinates, 
these  points  would  lie  on  a  line,  provided  the  substance  was  sub- 
jected only  to  pure  mechanical  adsorption. 

As  the  simplest  example  we  shall  choose  the  curves  and  data 
which  H.  Freundlich  *  derived  from  his  studies  of  the  adsorption 
of  certain  fatty  acids  by  charcoal.     Fig.  3  shows  the  adsorption  curve 


Fig.  3.    Adsorption  of  fatty  acids  by  charcoal.      (After  H.  Freundlich.) 

of  acetic  acid,  propionic  acid  and   succinic  acid,  in  direct  graphic 
representation  as  we  have  explained  on  page  22. 

They  are  derived  from  the  following  somewhat  abridged  data  of 
Freundlich: 


Acetic  acid. 
/  millimolsx  x  /    millimols    \ 


/  m  \gni.  charcoal/ 


0.0181 
0.0616 
0.2677 
0.8817 

2.785 


0.467 

0.801 

1.55 

2.48 

3.76 


Propionic  acid. 


/  millimols  \   x  /    millimols    \ 
A       cc.       /TO  Vgm.  charcoal/ 


0.0201 
0.0516 
0.2466 
0.6707 
1.580 


0.785 

1.22 

2.11 

2.94 

3.78 


Succinic  acid. 
/miIlimols\  x   I    millimols    \ 
V       cc.       /  m  \  gm.  charcoal/ 


0.0076 
0.0263 
0.0477 
0.2831 
1.161 


1.09 
1.70 
1.95 
3.26 
4.37 


24 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Fig.  4  shows  the  hne  passing  through  the  logarithms  of  these  data 
and  it  should  be  observed  that  all  logarithms  less  than  unity  are 
negative. 

The  tangents  to  the  angle  of  inclination  between  the  elements  of 


Fig.  4.     (After  H.  Freundlich.) 
the  curve   (acetic  acid,  propionic  acid  and  succinic  acid)  and  the 
abscissa  (log  c)  is  the  exponent  -.     The  distance  on  the  ordinate 

log  — )  from  the  zero  point  (origin  of  co-ordinates)  to  the  point  inter- 


secting the  uniting  lines  is  log  k.     They  have  these  values: 

Acetic  acid  -  =  0.425,     k  =  2.606. 

n 

Propionic  acid  -  =  0.354,     k  =  3.463. 

Succinic  acid  -  =  0.274,     k  =  4.426. 

n 

Since  the  values  observed  do  not  lie  all  in  the  same  line,  as  is  shown 
in  Fig.  4,  a  mean  value  for  the  angle  whose  tangent  is  n  may  be  de- 
rived by  means  of  a  protractor.  In  the  same  way,  log  k  is  not  de- 
rived from  the  intersection  of  the  last  element  of  the  curve,  but 
from  the  mean  value. 

The  exponent  -  conditions  the  shape  of  the  curve,  and  varies 
n 

within  moderate  limits.  Though  marked  exceptions  have  been  ob- 
served, it  fluctuates  usually  between  0.5  and  0.8  as  H.  Freundlich 
has  shown  in  his  numerous  experiments. 

The  constant  k  in  the  adsorption  formula  is,  in  an  ideal  case,  a 
natural  constant  which  may  be  as  characteristic  for  the  adsorbed 
substance  as  k  is  in  the  distribution  between  two  solvents. 

The  great  difficulty  lies  in  the  fact  that,  in  the  case  of  the  dispersed 
adsorbing  phase,  we  do  not  consider  the  mass,  which  may  be  easily 


SURFACES  25 

..<■ 

determined  by  weighing  or  measuring,  but  the  surface  which  may 
vary  greatly  with  the  selfsame  weight.  That  in  fact  not  the  mass 
but  the  surface  of  the  dispersed  phase  is  of  importance  in  adsorp- 
tion, is  evident  from  the  following. 

Freundlich  and  Schucht*  permitted  dyes  to  be  adsorbed  by 
amorphous,  i.e.,  colloidal,  mercuric  sulphid  flocks.  When  the  HgS 
became  crystallin  the  dye  dissolved  again.  We  have  here  a  counter- 
part of  enzym  action  in  which  the  adsorbed  enzym  (e.g.,  pepsin)  is 
liberated  when  the  adsorbing  substrate  (fibrin)  changes  its  surface 
as  it  is  split  up.  W.  Mecklenburg*  succeeded  in  obtaining  different 
curves  in  the  adsorption  of  phosphoric  acid  by  colloidal  stannic  acid 
and  of  arsenic  by  ferric  hydroxid  when,  starting  with  solutions  of 
identical  concentration,  he  precipitated  stannic  acid  and  ferric 
hydroxid  at  different  temperatures;  all  the  other  conditions  were 
identical.  The  lower  the  temperature  at  which  stannic  acid  or  ferric 
hydroxid  gel  were  formed  the  more  phosphoric  acid  or  arsenic  was 
adsorbed.  The  peculiarity  of  these  curves  was  the  similarity  in  their 
shape  which  is  described  by  mathematicians  as  "affinitive";  on  this 
account  Mecklenburg  called  them  affinitive  adsorption  curves. 
They  can  not  be  otherwise  explained  than  that  the  same  mass  of 
adsorbent  may  have  a  different  surface  depending  on  the  tempera- 
ture at  which  it  was  formed. 

Adsorption  is  a  phenomenon  which  is  conditioned  hy  the  decrease 
of  the  surface  tension  of  the  solvent  in  respect  to  the  dissolved  sub- 
stance, at  the  interface  between  the  solvent  and  adsorbent.  In 
1888,  G.  Quincke  showed  that  substances  which  decrease  the  surface 
tension  between  the  solvent  and  the  dispersed  phase  must  collect 
about  the  dispersed  phase  with  the  formation  of  a  film.  H.  Freund- 
lich has  elaborated  and  experimentally  established  the  theory  of 
adsorption  phenomena,  basing  his  ideas  on  Gibbs'  Theorem.^  The 
marked  diminution  of  the  surface  tension  of  water  by  fats,  fatty 
acid,  soaps,  albumin  and  its  cleavage  products,  and  enzymes  is 
characteristic;  and  it  is  not  surprising  that  these  substances  are 
very  easily  adsorbed. 

From  what  has  been  already  said,  adsorption  appears  to  be  a 
purely  physical  phenomenon  in  which  the  chemical  relations  between 
adsorbent  and  adsorbed  substance  play  no  part  whatever.     [This 

1  Gibbs'  Theorem  states:  A  dissolved  substance  is  positively  adsorbed  if  it 
depresses  the  surface  tension,  negatively  adsorbed  when  it  raises  it.  Willard 
Gibbs  deduced  these  relations  for  gaseous  mixtures  and  not  for  fluid  solutions. 
The  statement  of  W.  Gibbs,  that  a  small  amount  of  a  dissolved  substance  may 
powerfully  depress  surface  tension  but  cannot  raise  it  much,  is  likewise  important 
(for  further  proof  see  H.  Freundlich 's  "  KapiJlarchemie"). 


26  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

statement  is  too  dogmatic;  in  dealing  with  colloidal  particles  which 
approach  molecular  dimensions  no  sharp  Hne  can  be  drawn  between 
physical  and  chemical  forces.  Tr.]  We  shall,  therefore,  with  Wo. 
OsTWALD,  call  these  purely  physical  phenomena  mechanical  adsorip- 
tion. 

Most  of  the  investigations  on  adsorption  have  been  conducted 
with  pulverized  solids,  hydrophobe  colloids,  and  gels  as  adsorbents. 
From  a  biological  standpoint  studies  of  adsorption  by  hydrophile 
sols  are  of  especial  importance,  when  we  consider  for  example  what 
occurs  in  the  blood.  The  only  investigations  of  this  character  that  I 
am  acquainted  with  are  those  of  H.  Bechhold*^  on  the  distribution 
of  methylene  blue  between  water  and  serum  albumin.  The  volume 
in  solution  is  directly  obtained  by  ultrafiltration  and  the  amount  of 
methylene  blue  distributed  in  the  aqueous  filtrate  thus  obtained  is 
measured.  It  was  shown  that  in  very  dilute  solutions  of  methylene 
blue  the  major  part  is  held  firmly  by  the  albumin;  whereas  in  greater 
concentrations  the  distribution  is  displaced  in  the  direction  of  the 
water.  The  curve  is  similar  to  that  of  an  adsorption  curve.  [The 
work  of  A.  B.  Macallum,  "  Surface  Tension  and  Vital  Phenomena," 
University  of  Toronto  Studies.  Physiol.  Series  No.  8  (1912)  should 
be  read  in  this  connection  as  it  involves  the  adsorption  of  potassium 
and  its  concentration  at  surfaces.     Tr.] 

Based  on  what  has  been  said  hitherto,  we  might  believe  that 
nothing  is  easier  than  to  determine  by  the  curves  of  distribution 
whether  we  are  dealing  with  chemical  combination,  distribution 
between  two  solvents  or  adsorption.  If,  however,  we  examine  the 
experimental  data,  we  see  that  there  is  rarely  close  agreement 
between  observation  and  the  calculated  results. 

These  divergences  led  to  the  derivation  of  other  formulas  deter- 
mined by  the  following  considerations.  According  to  the  formula 
on  page  21  the  more  concentrated  a  solution,  the  more  is  there 
adsorbed  from  it.  Actually,  saturation  limits  are  reached  in  many 
cases.  This  may  be  explained  as  follows :  Each  interface  can  adsorb 
only  a  layer  of  a  definite  thickness,  and  saturation  is  reached  when 
this  layer  is  filled  with  adsorbed  molecules.  The  formulas  of 
Arrhenius,*  Rob  Marc,*  and  C.  G.  Schmidt*  were  made  to  be  in 
conformity  with  the  observed  facts. 

The  question,  whether  we  are  dealing  with  adsorption  or  solution, 
is  frequently  difficult  to  decide  when  the  dissolved  substance  has  a 
different  molecular  weight  in  the  dispersed  phase  than  in  the  solvent 
(see  p.  20).  The  curve  of  distribution  may  then  assume  the  form 
of  an  adsorption  curve  and  in  solving  the  problem  all  the  incidental 
circumstances  must  be  considered;  for  instance,  W.  Biltz*^  found 


SURFACES  27 

,.<■ 

that  the  distribution  of  arsenious  acid  between  iron  hydroxid  gel  and 
water  follows  the  equation 

X 

—  adsorbed 

3-  =  0.631. 

c  (free)5 

Were  we  to  believe  that  the  arsenious  acid  went  into  solid  solution  in 
iron  hydroxid  gel  we  should  have  to  conclude  that  the  arsenious  acid 
had  a  molecular  weight  one-fifth  as  large  in  iron  hydroxid  gel  as  in 
water.  From  other  observations,  however,  we  know  that  arsenious 
acid  in  water  breaks  up  into  simple  molecules  so  that  the  assumption 
that  it  dissolves  in  iron  hydroxid  gel  is  untenable. 

The  following  statements  will  show  that  many  conditions  may 
modify  the  course  of  the  adsorption  curve  and  such  cases,  according 
to  L.  Michaelis,  are  best  called  abnormal  adsorption. 

A  substance  may,  for  instance,  as  the  result  of  swelling,  absorb 
more  water  than  the  substance  dissolved  and  thereby  simulate  a  nega- 
tive adsorption.  Thus,  B.  Hekzog  and  Adler*  found  that  talcum 
powder  adsorbed  from  sugar  or  albumin  solution  more  water  than 
sugar  or  albumin  so  that  as  a  result,  the  solution  appeared  to  be  more 
concentrated  at  the  end  than  at  the  beginning  of  the  experiment. 

The  great  conceiitration  which  adsorbed  substances  cause  in  the 
dispersed  phase  may  be  associated  with  changes  in  its  condition; 
for  instance,  it  may  be  thrown  down  as  a  solid.  It  has  been  ob- 
served, that  charcoal  which  has  been  shaken  with  a  solution  of 
anilin  dye  shows  the  greenish  metallic  shimmer  and  the  dichroism 
of  the  solid  dye;  albumin  may  coagulate  at  interfaces.  Pro- 
found changes  may  accompany  these  variations  of  condition.  This 
may  be  shown  by  the  following  examples,  the  so-called  basic  anilin 
dyes  are  salts  consisting  of  a  strong  acid  (usually  hydrochloric 
acid)  and  a  weak  color  base.  The  aqueous  solution  undergoes 
strong  hydrolytic  dissociation;  the  free  color  base  shows  a  more  or 
less  colloidal  character  and  is  always  strongly  adsorbed.  H.  Freund- 
lich and  G.  Losev  *  showed  that  the  color  bases  adsorbed  by  char- 
coal from  new  f uchsin  and  crystal  violet  were  changed  at  the  surface 
of  the  charcoal,  and  substances  with  entirely  different  properties 
were  formed;  probably  they  were  the  insoluble  substances  which 
A.  VON  Baeyer  had  previously  prepared  separately.  In  the  case 
of  the  basic  dyes,  these  chemically  changed  substances  form  the  color 
on  the  textile  fiber. 

Now  we  must  recall  that  pure  adsorption  is  a  condition  of  equilib- 
rium changeable  with  the  concentration  of  the  dissolved  substance. 
There  can  be  no  balance  if  a  substance  is  removed  from  the  solution 


28  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

and  thus  made  insoluble  (irreversible)  as  in  the  above-mentioned  ex- 
ample of  the  basic  dyes.  As  dye  stuff  becomes  insoluble,  the  char- 
coal or  fibers  must  adsorb  more  dye  and  the  process  will  continue 
until  all  the  dye  is  adsorbed  from  the  solution.  In  the  case  in  ques- 
tion, the  process  is  prematurely  ended  because  of  the  concentration 
of  the  HCl  hydrolytically  separated,  which  to  a  certain  degree  ex- 
ercises a  solvent  action  on  the  colored  condensation  product.  Wilh. 
OsTWALD  *  has  called  attention  to  the  fact  that  in  solutions  which 
are  hydrolytically  split  into  fatty  acids  and  free  alkali,  there  is  a 
marked  displacement  of  the  adsorption  balance.  This  occurs  in 
washing.  The  fatty  acids  are  adsorbed  by  the  fabric  and  the  skin; 
to  accomplish  this,  there  must  occur  in  the  solution  a  further  hydroly- 
sis, i.e.,  a  splitting  off  of  alkali.  In  other  cases  the  dissolved  sub- 
stances may  be  completely  removed  from  solution,  e.g.,  albumin 
from  urine  (by  charcoal,  silicic  acid  or  mastic  emulsion,  as  adsorb- 
ents). 

If  we  try  to  determine  the  adsorption  curves  for  the  phenomena 
just  described  (especially  the  fixation  of  dyes),  we  shall  find  that  the 
curve  might  be  mistaken  for  that  of  an  irreversible  chemical  process. 

Still  more  peculiar  are  the  curves  which  W.  Biltz  and  H.  Steiner  * 
obtained  for  the  adsorption  of  night  blue  and  Victoria  blue  by 
cotton,  and  H.  Freundlich  *^  for  the  adsorption  of  strychnine  salts 
by  charcoal  or  arsenic  trisulphid,  as  well  as  G.  Dreyer  and  J. 
Sholto  *  for  the  adsorption  of  agglutinins  by  bacteria.  In  these 
cases  less  substance  was  taken  up  by  the  adsorbent  from  the  con- 
centrated solutions  than  from  those  of  medium  concentration. 

The  explanation  of  a  phenomenon  of  this  kind  was  given  by  A. 
Lottermoser,*  who  observed  with  A.  Rothe  that  amorphous  silver 
iodid  adsorbed  less  potassium  iodid  from  highly  concentrated  solu- 
tions than  from  solutions  of  medium  concentration.  The  process  is 
influenced  by  the  fact  that  high  concentrations  of  KI  precipitate 
Agl,  and  make  it  partially  assume  the  crystallin  form.  In  this  in- 
stance the  cause  is  a  diminution  of  surface;  in  the  other  cases,  above 
described,  there  is  a  strong  probability  of  a  diminution  of  surface, 
especially  with  the  agglutination  of  bacteria. 

Hitherto  it  has  been  tacitly  assumed  that  there  is  no  affinity  be- 
tween adsorbents  and  dissolved  substance.  This  occurs,  though  only 
in  a  few  exceptional  cases.  Thus  B.  S.  G.  Hedin  *^  has  shown  that 
certain  enzymes  (trypsin  and  rennin)  are  irreversibly  adsorbed  from 
water,  but  they  may  be  displaced  by  other  substances  (casein,  serum, 
grape  sugar).  L.  Michaelis  demonstrated  that  acid  kaolin  could 
adsorb  only  basic  or  amphoteric  dyes,  while  basic  clay  could  adsorb 
only  acid  dyes.     Similar  experiments  have  been  performed  by  various 


SURFACES  29 

..c 

other  experimenters  with  wool,  filter  paper,  etc.  Since  the  chemical 
constitution  of  many  of  these  substances  is  unknown  and  others  can- 
not be  prepared  free  of  electrolytes,  H.  Bechhold  considered  it  de- 
sirable to  settle  the  problem  by,  using  substances  possessing  definite 
constitution  and  which  were  easily  obtained  in  a  pure  state.  As  such 
adsorbents,  he  chose  naphthalin  (CioHs,  neutral),  naphthol  (C10H7OH, 
acid),naphthylamin(CioH7NH2,  basic),  amidonaphthol  (C10H6OHNH2, 
amphoteric).  These  substances  were  freely  suspended  in  water  and 
shaken  for  several  minutes  with  solutions  of  various  dyes;  they  were 
filtered  off  and  washed  until  the  filtrate  was  practically  colorless. 
The  dye  solutions  employed  were  those  generally  used  in  microscopic 
technic.  The  results  of  my  staining  experiments  are  shown  in  the 
accompanying  table.  From  these  experiments  it  is  evident  that  in 
most  cases,  even  with  the  neutral  naphthalin,  there  is  at  least  a  faint 
staining.  The  coloration  is  so  slight,  that  it  certainly  could  not  be 
recognized  in  a  microscopic  specimen  and  it  is  evidently  to  be  attrib- 
uted to  mechanical  adsorption.  It  may  be  seen  at  a  glance  how  re- 
markably the  staining  differs,  depending  on  the  chemical  constitution 
of  the  stained  substance,  neutral  naphthalin  is  not  deeply  colored 
by  any  stain.  Naphthylamin  and  amidonaphthol  are  always  most 
strongly  stained  by  the  acid  dyes,  and  naphthol  and  amidonaphthol 
by  the  color  bases.  Thus  we  see  that  the  chemical  constitution  of  the 
adsorbent  plays  a  very  important  part  in  the  distribution  of  the 
dissolved  substance  between  the  solvent  and  the  dispersed  phase. 
Berczeller  and  Czaki*  reached  analogous  results  with  the  adsorp- 
tion of  alkaloids  (cocain,  atropin,  etc.),  by  various  powders  (starches, 
coagulated  albumin,  which  acted  as  weak  acids  and  adsorbed  most 
strongly,  whereas  alkaline  CaCOs  adsorbed  least) .  That  we  are  deal- 
ing with  electro che7nical  phenomena  in  the  above  cases  is  still  more 
evident  when  we  observe  how  the  addition  of  electrolytes  affects  adsorb- 
abihty.  Wool,  which  is  dyed  particularly  well  by  basic  dyes  in  a 
neutral  bath,  takes  them  up  still  better  from  an  alkahne  bath,  but  it 
is  also  dyed  in  an  acid  bath  with  acid  colors.  Still  better  evidence 
lies  in  the  fact  that  the  cations  of  neutral  salts  increase  the  dyeing 
of  acid  colors  in  proportion  to  the  valence  of  the  cation  (W.  M. 
Bayliss). 

We  must  furthermore  mention  the  fact  that  supplementary  chemical 
reactions  may  occur  between  adsorbent  and  adsorbed  substances, 
which  may  lead  to  a  fixation  that  makes  the  process  irreversible,  i.e., 
a  true  chemical  combination  may  result.  The  occurrence  of  this 
condition  is  characterized  by  the  fact  that  it  requires  a  certain 
length  of  time,  whereas  according  to  H.  Freundlich,  the  adsorp- 
tion balance  is  established  in  a  few  minutes.     Moreover,  such  a  de- 


30 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


1 

'^midonaphthol 

Naphthalin. 

/3-Naphthol.       /3-Naphthylamin. 

(freshly  precipi- 
tated). 

Basic  dyes: 

Methylene  blue 

very  faint 
blue 

dark  blue 

very  faint 
blue 

LöfHer's  blue  ^ 

faint  blue 

dark  blue 

almost  un- 
colored 

blue 

Carbol-fuchsin 

faint  red 

reddish 

almost  un- 

colored 
faint  blue 

deep  red 

Crystal  violet 

bluish 

deep  blue 

deep  violet 

blue 

Bismark  brown 

iiiTiit  fi  iTipn 

brownish 

practically 
unstained 

LlllOLcmiCVJ. 

Gram's  Stain: 

Anilin  water  —  gen- 

light violet 

deep  violet 

faint  violet 

deep  violet 

tian  violet 

Treatment  with  io- 

almost  com- 

din-potassium io- 

pletely  de- 

did solution. 

colorized 

dark  blue 

blue 

dark  blue 

Washing  off  with  al- 

almost com- 

undeter- 

completely 

deep  violet 

cohol.                        !- 

pletely  de- 

mined be- 

decolor- 

(as far  as 

colorized 

cause   too 
soluble 

ized 

could    be 
deter- 
mined  in 
view   of 
great 
solubil- 

' 

ity) 

Acid  dyes: 

Eosin 

unstained 
faintly  yel- 

unstained 
very   faintly 

pink 
reddish  yel- 

red 

Aurantia 

yellow 

low 

yellow 

low 

Picric  acid 

unstained 

almost  un- 
stained 

very  faint 
yellow 

almost    un- 

stained 

Alizarin  (dissolved  in 

brownish 

brownish 

violet 

violet 

KOH) 

violet 

Gallein 

unstained 

almost  un- 
stained 

almost  un- 
stained 

faint  brown- 

ish red 

Chrome  violet 

almost  un- 

almost un- 

almost un- 

red 

stained 

stained 

stained 

Amphoteric  dyes: 

Benzopurpurin 

reddish 

reddish 
(somewhat 
deep  er 
than    the 
other  s 
and     mot- 
tled blue) 

reddish 

red 

Janus  red 

reddish 

deep  red 

reddish 

deep  red 

Mixtures: 

Triacid 

faint    bluish 

deep   bluish 

very  faint 

dark  bluish 

green 

green 

violet 

green 

1  Löffler's  blue  is  a  methylene  blue  solution  with  a  trace  of  alkali;  Carbol-fuchsin  is  a  fuchsin  solu- 
tion containing  phenol.  Both  of  these  as  well  as  Gram's  stain  are  employed  in  staining  bacteria. 
The  latter  affords  an  opportunity  for  the  differentiation  of  certain  bacteria  and  cocci.  Although 
many  bacteria  are  entirely  or  partly  decolorized  by  subsequent  treatment  with  the  iodin-potassium 
iodid  solution  and  washing  with  alcohol,  many  retain  their  stain. 


SURFACES  31 

layed  supplementary  process  may  indicate  a  slow  diffusion  of  the 
adsorbed  substance  into  the  adsorbent,  as  J.  Davis  has  demon- 
strated in  the  case  of  the  adsorption  of  iodin  by  charcoal.  In  the 
fixation  of  dyes  by  textile  fibers  we  can  assume  the  probable  occur- 
rence of  secondary  chemical  processes.  I  believe  that  many  mis- 
understandings in  the  moot  questions  of  toxin-antitoxin  fixation  might 
have  been  avoided,  if  there  had  been  a  clear  understanding  of  the 
various  phenomena  which  may  occur  in  the  course  of  an  adsorption 
process. 

Finally,  it  must  be  mentioned  that  catalyzers  are  adsorbed  (all 
organic  ferments  are  colloids).  By  reason  of  adsorption,  the  reaction 
in  a  solution  may  be  stopped;  or  in  other  cases,  the  reaction  may  be 
accelerated  upon  the  adsorbent.  Thus  oxidations  may  be  brought 
about  by  concentration  of  oxygen  on  the  adsorbent  or  reductions  by 
concentration  of  hydrogen  (C.  Paals'  reduction  of  nitrobenzol  by 
colloidal  palladium  and  other  chemical  reactions.) 

The  fundamental  conception  of  adsorption  is  so  illuminating  that 
the  attempt  has  been  made  to  explain  a  large  series  of  biological 
processes  as  adsorption  phenomena  (enzym  action,  union  of  toxin  and 
antitoxin,  etc.).  I  cannot  better  express  the  results  of  all  this  work 
than  in  the  words  of  W.  Biltz,*'*  who^  in  another  connection,  says, 
"  The  testing  ...  of  the  material  in  accordance  with  an  exact 
method,  such  as  is  involved  in  the  use  of  a  formula,  offers  the  worker 
a  rather  mingled  pleasure,  as  may  be  noted  from  the  great  difference 
between  the  results  of  experiment  and  of  calculation.  If  it  were  not 
for  the  novelty  of  the  subject  investigated,  .  .  .  the  result  which  is 
so  frequently  accepted  for  the  sake  of  the  principle,  would  be  of  very 
little  importance." 

I  should  like  to  add  further:  The  adsorption  formula  is  a  rock 
which,  Lorelei-like,  has  magically  attracted  numberless  scientific 
voyagers  only  to  wreck  many  of  them.  Every  complicated  phe- 
nomenon of  higher  organisms,  which  consists  partly  of  chemical, 
partly  of  solution  phenomena  and  perhaps  partly  of  true  adsorption, 
must  assume  the  general  character  of  an  adsorption  and  have  a 
formula  which  seems  a  cross  between  a  chemical  process  and  a  solu- 
tion. Thus,  if  a  biological  phenomenon  seems  to  suit  the  formula  of 
an  adsorption,  this  may  be  merely  a  sign  post,  which,  alas,  many  an 
investigator  may  mistake  for  the  goal. 

What  then  is  the  biological  significance  of  what  we  have  distinguished 
as  chemical  combination,  solution  and  adsorption?  Here  we  approach 
the  most  important  principle  governing  the  processes  in  the  living 
organism;  it  is  what  P.  Ehrlich  describes  as  distribution.  The  de- 
veloping and  the  fully  developed  organism  are  constantly  receiving 


32 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


food  materials,  which  are  taken  up  at  the  places  where  needed,  stored, 
and  when  necessary  given  up  again.  In  other  words,  the  organ- 
ism, plant  as  well  as  animal,  is  a  vessel  containing  an  aqueous  solu- 
tion in  which  various  colloids  exist  as  dispersed  phases.  The  balance 
which  rules  at  each  moment  is  disturbed  by  food  material  entering 
the  vessel  and  by  the  metabolic  products  developing  in  it,  and 
these  become  distributed  between  the  solvent  and  the  dispersed 
phase,  the  organ-colloids.  In  this  entire  chapter  we  have  treated 
the  simple  case  occurring  when  a  single  dispersed  phase  exists  in  a 
single  dispersing  medium  (solvent).  We  may  still  assume  without 
serious  error,  that  there  is  one  dispersing  medium;  but  instead  of 
one  dispersed  phase  in  the  organism  we  have  dozens,  perhaps  hun- 
dreds, of  dispersed  phases.  Each  individual  class  of  cells  is  a  dif- 
ferent dispersed  phase  with  different  properties.  Only  in  this  way 
can  we  understand  how  the  assimilation  products  are  sorted,  stored 
up  and  changed  into  the  tissues  of  the  several  organs. 

To  quote  a  single  example,  we  find  (according  to  a  table  in  E. 
Abderhalden's  Textbook  of  Physiological  Chemistry)  that  there  are 
the  following  distributions: 


to  1000  parts  by  weight 

Serum  of  ox. 

Blood  corpus- 
cles of  ox. 

Sugar 

Cholesterin 

Lecithin 

Soda 

Potash 

Lime 

Ferric  oxid 

1.05 

1.238 

1.675 

4.312 

0.255 

0.119 

3^379 
3.748 
2.232 
0.722 

1^671 

It  is  difficult  to  imagine  a  more  unequal  distribution.  For  in- 
stance, potassium  and  sodium  salts  enter  the  circulation  as  electro- 
lytes to  the  same  extent;  and  there  can  be  no  question  of  any 
irreversible  chemical  combination  of  these  salts  either  with  the  serum 
or  with  the  blood  corpuscles,  since  both  diffuse  away  when  the  serum 
or  blood  corpuscles  are  brought  in  contact  with  pure  water.  There 
is  thus  a  condition  of  equilibrium  in  the  blood  by  which  the  hlood 
corpuscles  dissolve  or  adsorb  proportionately  more  potassium  salts, 
while  the  serum  albumin  dissolves  or  adsorbs  more  sodium  salts.  This 
is  not  a  unique  case,  for  humus  adsorbs  chiefly  the  potassium  salts 
from  a  mixture  and  permits  the  sodium  salts  to  pass  through.  With 
iron  the  conditions  are  different;  it  certainly  must  enter  the  organism 


SURFACES  33 

in  some  mobile  form,  yet  at  the  places  where  blood  corpuscles  are 
formed  it  is  chemically  fixed  as  hemoglobin.  We  may  say  d  priori 
that  the  products  of  dissimilation  become  very  soluble  in  the  dispers- 
ing medium,  being  dissolved  or  adsorbed  but  slightly  by  the  dispersed 
phase;  in  fact  they  are  not  chemically  fixed  at  all,  so  that  they  leave 
the  body  chiefly  in  the  urine;  they  are,  indeed,  crystalloids  of  which 
only  so  much  is  retained  in  the  blood  by  solution  or  adsorption  as  is 
necessary  for  a  proper  balance. 

We  must  here  curtail  our  remarks  and  refer  the  reader  to  the 
chapter  on  Distribution  of  Substances  and  Metabolism. 

What  applies  to  the  substances  necessary  for  the  maintenance  of 
the  organism  applies  also  to  such  foreign  substances  as  have  a  toxic 
or  pharmacodynamic  effect.  It  is  a  principle  that  such  foreign 
bodies  as  are  chemically  fixed,  permanently  injure  the  affected  cell; 
narcosis  seems  to  me,  to  be  a  typical  example  of  simple  solution, 
a  process  that  is  completely  reversible.  [Permanent  injury  may  also 
be  caused  by  the  breaking  of  an  emulsion.  See  M.  H.  Fischer  and 
Marian  O.  Hooker,  "  Fats  and  Fatty  Degeneration,"  John  Wiley  & 
Sons,  New  York,  1917.  Tr.]  Between  these  extreme  cases  there  are 
substances  which  are  adsorbed,  and  are  active  even  in  small  doses, 
though  larger  doses  do  not  cause  materially  greater  damage.  Under 
favorable  conditions  these  processes  may  be  reversible.  The  details 
are  treated  of  in  the  chapter  on  Toxicology  and  Pharmacology. 

Finally,  I  wish  to  refer  to  the  chapter  on  Immunity  Reactions 
where  it  is  an  important  question  whether  chemical  combination, 
solution  or  adsorption  obtains. 

Surface  Skins. 

Absolutely  pure  water  has  no  surface  viscidity,  which  means  that 
a  metal  or  glass  disc  suspended  by  a  thread  at  the  surface  performs 
as  many  oscillations  after  a  single  turn  as  it  does  when  immersed. 
The  slightest  impurities  may,  however,  suffice  to  cause  a  marked 
retarding  effect  at  the  surface.  On  page  25  we  saw  that  substances 
which  lower  the  surface  tension  of  a  fluid  concentrate  and  spread 
out  at  the  surface,  so  it  is  to  be  expected  that  the  surface  will  have 
a  different  viscidity  from  the  interior.  Poggendorff  and  Plateau 
were  the  first  to  study  the  formation  of  skins  on  fluids,  but  we  are 
indebted  to  M.  V.  Metcalf,*  G.  Nagel  *  and  E.  Rohde  *  for  recent 
studies  that  have  clarified  the  subject.  It  was  shown  as  the  result 
of  these  investigations  that  colloids  and  substances  at  the  border  line 
between  colloids  and  crystalloids,  especially  many  dyes,  as  fuchsin, 
methyl  violet,  peptones  and  several  other  substances,  concentrate  at 


34  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

the  surface  of  aqueous  solutions  and  form  a  layer,  which  at  first  is 
easily  movable.  In  a  short  time  there  is  a  change  in  this  layer.  In 
the  case  of  staining  solutions  the  surface  appears  dull  after  an  hour 
and  gradually  there  is  formed  a  solid  layer  which  histologists  and 
bacteriologists  know  to  their  sorrow.  It  is,  therefore,  necessary  to 
filter  aqueous  solutions  of  stains  each  time  before  using,  even  though 
they  have  been  protected  from  dust.  The  dye  concentrated  at 
the  surface  undergoes  chemical  changes  which  the  investigations 
showed  to  be  independent  of  the  gas  upon  the  surface  (it  might  have 
been  attributed  to  oxidations  with  oxygen  or  to  CO2,  etc.).  What 
has  been  said  of  dye  solutions  occurs  also  in  the  case  of  peptone 
solutions,  as  Metcalf  demonstrated. 

The  thickness  of  the  layer  which  will  just  form  a  solid  skin  has 
been  measured,  and  found  to  be,  for  peptone  3  fxß  (Metcalf),  for  albu- 
min 3  to  7  jU;U  (Devaux).  Thus  it  is  probably  many  times  greater 
than  the  hypothetical  diameter  of  a  molecule,  perhaps  even  equaling 
the  radius  of  molecular  attraction.  The  process  of  skin  formation 
may  be  very  much  hastened  by  amplifying  the  surface,  i.e.,  by  shak- 
ing the  fluid  or  passing  gas  through  it.  Thus  W.  Ramsden  *  was 
able  to  remove  by  shaking  practically  all  the  albumin  from  an 
albumin  solution.  The  albumin  passed  into  the  foam  and  there 
formed  solid  skins.  This  phenomenon  is  of  greatest  practical  impor- 
tance, since  the  solidity  of  meringue,  whipped  cream  and  beer  foam 
evidently  depends  upon  it.  In  the  case  of  beer,  the  rising  bubbles  of 
CO2  carry  foam-forming  colloids  at  their  surfaces  and  conversely  the 
beer  foam  exerts  a  tension  (pressure)  which  hinders  the  escape  of 
CO2  and  thus  keeps  the  beer  fresh  for  a  longer  time.  Everyone  who 
has  worked  with  colloidal  solutions  knows  how  high  the  gas  pressure 
must  be  in  order  that  a  stream  of  gas  may  be  forced  through  a  solu- 
tion which  has  once  formed  a  layer  of  foam.  The  formation  of  a 
skin  on  boiling  milk  is  evidently  to  be  classed  among  these  phe- 
nomena. The  significance  of  this  process  for  the  coagulation  of  fibrin 
has  been  indicated  on  page  299. 

To  these  phenomena  belongs  'Hnadivation  of  ferments  hy  shaking" 
described  by  E.  Abderhalden  and  Guggenheim  *  and  independ- 
ently by  Signe  and  Sigval  Schmidt-Nielsen.* 

The  formation  of  surface  skins  is  so  sharply  characteristic,  that 
mixtures  of  substances,  which  diminish  the  surface  tension  of  water 
to  different  degrees,  may  be  separated  by  shaking.  Until  now,  we 
have  had  only  the  qualitative  investigations  of  W.  Ramsden,*  who 
determined  the  predominance  of  saponin  in  the  foam  on  shaking  a 
mixture  of  saponin  and  albumin.  (Saponin  lowers  the  surface 
tension  of  water  more  than  albumin.) 


SURFACES  35 

Hitherto  we  have  only  regarded  the  formation  of  surface  skins  at 
the  interface  fluid/gas.  Such  skins  may  be  formed,  however,  at 
the  interface  fluid/fluid  or  fluid/solid,  provided  only  that  the  sub- 
stance in  question  diminishes  the  surface  tension  of  the  water  with 
reference  to  the  other  fluid  or  the  solid  phase.  J.  Ziegler  has  in- 
formed me  (in  a  private  communication)  that  on  shaking  benzol, 
toluol,  etc.,  with  water  containing  albumin  or  gelatin,  the  benzol 
or  toluol  forms  above  the  water  an  emulsion  which  contains  the 
colloid,  and  that  with  repeated  shaking,  the  major  part  of  the 
colloid  may  be  removed  from  the  aqueous  solution.  Shortly  after 
this  communication,  there  appeared  a  publication  by  Winkelblech  * 
which  not  only  confirmed  these  facts  but  called  attention  to  the  fact 
that  through  the  formation  of  an  emulsion  mere  traces  of  colloids 
could  be  detected.  This  phenomenon  has  long  been  recognized  as  a 
very  disturbing  factor  by  organic  chemists.  On  shaking  reaction 
mixtures  with  ether  or  benzol,  such  emulsions  frequently  form  and 
are  very  difficult  to  separate.  We  know  now  that  these  emulsions 
are  to  be  attributed  to  the  formation  of  colloidal  reaction  products. 

H.  Bechhold  and  J.  Ziegler  used  the  method  of  shaking  out 
the  foam  for  the  separation  of  albumoses  (Witte's  peptone)  into 
their  components.  They  shook  a  10  per  cent  aqueous  solution  of 
Witte's  peptone  with  ether,  separated  the  ether  foam  from  the 
aqueous  solution  and  again  shook  it  out  with  ether.  Then  the 
ether  was  permitted  to  evaporate  from  the  foam,  and  the  residue  was 
dissolved  in  ten  times  its  volume  of  water,  and  this  solution  was 
again  shaken  out  with  ether.  After  thus  treating  the  solution  from 
three  to  five  times,  two  substances  were  obtained,  one  of  which  re- 
mained in  the  water  in  clear  solution  and  became  turbid  when  treated 
with  24  to  25  per  cent  of  ammonium  sulphate.  By  this  procedure  a 
separation  of  two  components  was  obtained,  but  it  is  a  question 
whether  the  water-insoluble  portion  was  present  in  the  original  solu- 
tions or  was  formed  by  the  shaking,  like  Metcalf's  peptone  skins. 
The  slight  diminution  in  concentration  is  in  favor  of  the  first  view. 
By  "shaking  out  the  foam  "  a  separation  of  the  slightly  water-soluble 
hetero-albumoses  and  the  remaining  albumoses  was  accomplished. 

This  spreading  of  colloids  and  the  formation  of  colloid  films  at 
the  interface  between  two  fluids  is  a  phenomenon  very  frequently 
observed.  G.  Quincke  *^  showed  that  gum  coflects  at  the  interface 
between  oil  and  a  gum  arable  solution.  Pharmaceutical  emulsions 
accordingly  consist  of '  oil  spheres  surrounded  by  a  film  of  gum. 
And  when  oil  is  emulsified  with  albumin,  oil  spheres  surrounded  by  a 
film  of  albumin  are  formed  (Ascherson  *).  I  have  attributed  the 
so-called  ''serum  films  surrounding  the  globules  in  milk"  to  the  for- 


36  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

mation  of  surface  skins,  see  page  346.  Where  oil  globules  occur  in 
aqueous  solutions  containing  colloids,  it  may  be  assumed  that  they 
are  surrounded  by  a  film  of  colloid  which  prevents  them  from  run- 
ning together  and  forming  larger  drops  of  fat.  This  is  exemplified  in 
the  emulsion  of  fats  in  the  intestine  and  in  the  milky  turbidity  of  the 
serum  after  ingestion  of  fat  as  well  as  in  the  oleaginous  and  resinous 
emulsions  in  plants,  e.g.,  in  the  milky  sap  of  the  Euphorbiace. 

As  has  been  already  stated  the  same  conditions  obtain  for  the 
interfaces  between  fluid/solid  as  for  fluid/fluid.  H,  Bechhold*^ 
explains  the  action  of  protective  colloids  (see  p.  11)  as  a  mani- 
festation of  this  phenomenon.  Protective  colloids  form  colloidal 
films  about  the  substance  in  suspension  and  thus  impede  the 
coalescence  (flocculation)  of  the  separate  particles.  Consequently 
surface  pellicles  afford  stability  to  the  metal  sols,  and  permit  their 
practical  utilization. 

On  the  other  hand,  suspensions  and  hydrophobe  colloids,  depend- 
ing on  circumstances  and  the  surface  tension,  may  either  pass  from 
one  fluid  into  another  fluid  with  which  it  is  not  miscible,  or  they  may 
concentrate  at  the  interface  (Reinders).  This  must  be  considered 
in  all  studies  on  the  distribution  of  colloids  in  the  body,  as  in  staining 
with  colloidal  dyes  and  the  injection  of  colloidal  metals  and  possibly 
even  infection  with  micro-organisms. 

[According  to  the  views  of  Martin  H.  Fischer  and  Marian  0. 
Hooker,^  we  must  distinguish  between  the  making  of  an  emulsion 

^  See  Martin  H.  Fischer  and  Marian  O.  Hooker,  Fats  and  Fatty  Degen- 
eration, John  Wiley  and  Sons,  New  York,  1917,  where  references  to  their  earher 
pubhcations  may  be  found. 

"  Both  W.  D.  Bancroft  and  G.  H.  A.  Clowes  at  the  Urbana  (1916)  meeting 
of  the  American  Chemical  Society,  in  their  discussion  of  our  own  views  regarding 
the  importance  of  colloid  solvates  (colloid  hydrates)  for  the  stabilization  of  emul- 
sions, found  in  om-  views  something  irreconcilable  with  their  notions  of  the  im- 
portance of  interfacial  films  and  of  surface  tension  changes  in  these.  While  we 
do  not  wish  to  insist  upon  a  harmony  where  such  may  not  be  desired,  there  is,  of 
course,  nothing  mutually  exclusive  in  the  ideas  of  solvation,  of  changes  in  surface 
tension,  and  —  at  times  —  the  formation  of  a  continuous  third  phase  between 
the  two  chief  substances  making  up  an  emulsion.  When  "water,"  according  to 
our  notion,  becomes  a  "colloid  hydrate,"  the  properties  of  the  second  liquid  are 
different  from  those  of  the  first,  and  these  properties  include  surface  tension, 
viscosity  and  distribution  between  two  phases.  But,  we  repeat,  these  factors  to 
which  Plateau,  Quincke  and  Pickering  first  directed  attention  are  not  by  them- 
selves able  to  explain  all  the  phenomena  observed.  Where  Clowes  holds  that  an 
emulsion  of  oil  is  stabilized  through  sodium  oleate  because  the  substance  reduces 
the  surface  tension  of  water,  we  would  say  that  stabilization  has  ensued  because 
the  oil  has  been  divided  into  a  highly  hydratable  sodium  soap.  When  the  addi- 
tion of  calcium  destroys  this  emulsion,  it  is  not  because  of  complicated  changes  in 
a  surface  film,  but  simply  because  calcium  oleate  is  an  only  slightly  hydratable 


SURFACES  37 

.J 

and  its  stabilization.  The  making  of  an  emulsion  is  essentially  a 
mechanical  process  concerned  with  the  mere  obtaining  of  the  sub- 
division of  one  liquid  in  a  second,  as  oil  in  water.  The  problem  of 
the  stabilization,  after  such  a  subdivision  has  been  brought  about, 
is  a  totally  different  matter.  In  a  certain  sense  the  main  feature  in 
this  stabihzation  consists  of  the  getting  rid  of  the  water  as  such  in  the 
emulsion  and  substituting  for  it  a  hydrated  colloid. 

An  emulsion  is  stabilized  through  any  so-called  emulsifying  agent 
only  because  this  emulsifying  agent  is  a  hydrophilic  (lyophilic)  col- 
loid. Oil,  for  example,  cannot  be  permanently  emulsified  in  water 
in  amounts  exceeding  a  fraction  of  one  per  cent,  but  in  a  medium  in 
which  the  water  is  bound  to  an  emulsifying  agent  as  a  hydrate  (or 
solvate)  the  oil  content  can  be  carried  to  a  very  high  figure  (50  or  60 
per  cent) .  When,  for  example,  acacia  is  used  as  an  emulsifying  agent, 
it  means  that  the  permanent  emulsion  is  made  permanent  because 
the  acacia  unites  with  the  water  to  form  an  acacia-hydrate. 

After  the  stabilization  of  an  emulsion  has  been  accomplished 
through  the  production  of  a  colloid  hydrate,  secondary  concentration 
effects  may  be  brought  about  which  lead  to  a  concentration  of  the 
colloid  material  upon  or  in  the  surface  of  the  oil  droplets  but  these 
secondary  effects  are  not  to  be  confused  with  the  primary  ones  neces- 
sary for  the  stabilization  of  the  emulsion. 

W.  D.  Bancroft  asserts  in  a  review  of  Fischer's  book  that  there 
are  no  criteria  which  these  alleged  compounds  (the  solvates)  could 
satisfy.     J.  of  Ind.  &  Eng.  Gh.,  vol.  9,  No.  12,  Dec,  1917. 

Bancroft  observed  that  while  soaps  of  mono-valent  cations  used 
as  emulsifying  agents  for  oil  and  water,  promote  the  formation  of 
emulsions  like  cream,  in  which  oil  is  dispersed  in  a  continuous  water 
phase,  soaps  of  di-  and  tri-valent  cations  form  emulsions  of  the 
opposite  type  like  butter,  in  which  water  is  dispersed  in  oil.  Ban- 
croft considers  that  soaps  of  sodium  or  potassium,  being  readily  dis- 
persed in  water  but  not  in  oil,  form  an  interfacial  film  or  membrane, 
the  surface  tension  on  the  water  side  of  which  is  much  lower  than  on 
the  oil,  and  that  consequently  an  emulsion  of  oil  in  water  is  formed, 

soap.  Free  water,  in  consequence,  appears  in  the  mixture,  and  the  oü  separates 
out  in  gross  form,  as  described  above,  for  only  very  Httle  oil  can  be  permanently 
subdivided  in  "pure"  or  "free"  water.  We  describe  the  consequences  of  such 
changes  from  highly  hydratable  to  less  hydratable  soaps  upon  the  stabüity  of  an 
emulsion  on  p.  49. 

Neither  do  we  wish  our  statement  that  an  agreement  is  possible  between 
Clowes'  and  our  views  on  simple  emulsions  to  be  expanded  to  include  his  behefs 
regarding  the  biological  behavior  of  the  fat  in  living  cells.  We  long  ago  gave  up 
the  notion  of  lipoid  membranes  about  cells  and  the  complex  notions  of  their 
changing  permeability  to  which  Clowes  and  many  authors  still  hold." 


38 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


1      ^/HzO 

H20 

^4  HgO 

oilL 

biL 

Emulsifying    Na  oleate  alone 

agent  or  Na  oleate  in 

or  film  excess  of  Ca 

oleate  equiv 

Emulsion 

formed  Oil  in  water 


Equiv.  chem. 

proportion 

Na  oleate  and 

Ca  oleate 


Ca  oleate  alone 
or  Ca  oleate  in 
excess  of  Na 
oleate  equiv. 


Critical  point         Water  in  oil 


Fig.  4a.     Conversion  of  oil-water  to 
water-oil  emulsion. 


while  soaps  of  calcium  and  magnesium,  being  readily  dispersed  in  oil 
but  not  in  water,  form  a  film,  the  surface  tension  on  the  oil  side  of 
which  is  lower  than  on  the  water,  and  consequently  an  emulsion  of 
water  in  oil  is  produced. 

Clowes  showed  that  emulsions  of  oil  in  water  could  be  converted 
into  emulsions  of  water  in  oil  and  vice  versa  by  varying  the  propor- 
tions of  certain  electrolytes 
added  to  the  system.  When 
equal  volumes  of  oil  contain- 
ing fatty  acid  and  water  con- 
taining NaOH  were  shaken 
together  sodium  soap  was  pro- 
duced and  an  emulsion  of  oil 
in  water  formed.  On  shaking 
this  emulsion  with  increasing 
proportions  of  calcium  chlor- 
id,  a  critical  point  at  which 
oil  and  water  separated  into 
two  distinct  layers  was  ob- 
served when  the  CaCl2  was 
added  in  sufficient  amount  to  convert  half  the  sodium  soap  into  cal- 
cium soap.  Further  addition  of  calcium  chlorid  led  to  the  forma- 
tion of  a  stable  emulsion  of  water  dispersed  in  oil.  Conversely,  the 
latter  emulsion  could  be  reconverted  through  the  critical  point  into 
one  of  oil  in  water  by  shaking  with  the  requisite  proportions  of  sodium 
soap  or  caustic  soda.     (See  diagram.  Fig.  4a.) 

Clowes  attributes  these  transformations  to  variations  in  the  sur- 
face tension  relations  of  the  water  and  oil  phases,  caused  by  varia- 
tions in  the  proportions  of  the  hydrophilic  sodium  soaps  which  lowers 
the  surface  tension  of  the  water  phase,  and  the  lipophilic  calcium  soap 
which  lowers  the  surface  tension  of  the  oil  phase.  An  emulsion  of  oil 
in  water  is  produced  when  the  surface  tension  is  lower  on  the  water 
than  on  the  oil  side  of  the  stabiUzing  film  or  membrane  formed  by 
concentration  of  the  emulsifying  agent  at  the  oil-water  interface.  A 
critical  point  occurs  when  the  surface  tension  is  equal  or  compensa- 
tory on  both  sides  of  the  film,  and  an  emulsion  of  water  in  oil  is  formed 
when  the  surface  tension  is  lower  on  the  oil  side  than  on  the  water 
side. 

Electrolytes  appear  to  exert  a  marked  effect  on  emulsion  equilib- 
rium, those  having  a  more  reactive  or  more  readily  adsorbed  anion 
appear  to  promote  the  formation  of  emulsions  of  oil  in  water,  while 
those  having  a  more  reactive  or  readily  adsorbed  cation  exert  the 
reverse  effect,  promoting  the  formation  of  emulsions  of  water  in  oil. 


SURFACES  39 

.J 

The  antagonistic  effects  exerted  by  electrolytes  of  these  two  opposing 

groups  appear  to  correspond  sufficiently  closely  with  those  observed 

by  OsTERHOUT  in  experiments  on  Hving  cells  to  suggest  the  possibihty 

that  variations  in  permeabihty  exhibited  by  protoplasm  under  the 

influence  of  various  salts  might  be  attributable  at  least  in  part  to 

reversible  transformations  of  the  marginal  layer  of  protoplasmic 

material  between  systems  in  which  a  non-aqueous  phase  is  dispersed 

in  an  aqueous,  which  would  be  relatively  freely  permeable  to  water, 

and  the  reverse  type  of  system  in  which  an  aqueous  phase  is  more  or 

less  surrounded  by  a  non-aqueous  film  which  would  be  impermeable 

or  relatively  less  permeable  to  water. 

A  Traube  capillary  pipette  was  employed  to  study  the  influence 
exerted  by  given  salts  individually,  and  in  combination,  on  the  rela- 
tive degree  of  dispersion  of  interfacial  soap  films  in  oil  and  water. 
Aqueous  solutions  containing  caustic  soda  or  soap  and  various  con- 
centrations of  the  salts  to  be  tested  were  allowed  to  flow  from  the 
capillary  pipette  through  neutral  oil  or  oil  containing  free  fatty  acid, 
and  the  number  of  drops  produced  served  as  an  index  of  the  dis- 
persing or  protective  effect  exerted  by  the  electrolytes  in  question 
on  the  interfacial  soap  film.  Those  electrolytes  which  possess  a 
readily  adsorbed  anion  appear  to  cause  an  increase  in  the  number  of 
drops,  which  corresponds  with  a  lowering  of  the  surface  tension  of 
the  water  phase,  a  destruction  of  the  surface  film,  and  an  increased 
permeability  of  the  system  to  water.  Those  electrolytes  which 
possess  a  readily  adsorbed  cation  exert  the  reverse  effect,  diminish- 
ing the  number  of  drops,  which  indicates  diminished  dispersion  or 
destruction  of  the  film  and  a  diminished  permeability  of  the  system 
to  water.  For  example,  a  0.001-m  NaOH  passed  through  ohve  oil 
gave  44  drops;  the  addition  of  NaCl  to  a  concentration  of  0.15-?7i 
raised  the  number  of  drops  to  300;  the  addition  of  CaCl2  at  a  concen- 
tration of  0.0015-w  lowered  the  number  of  drops  to  24;  while  a 
system  in  which  O.OOl-w  NaOH  was  employed  in  conjunction  with 
O.lS^m,  NaCl  and  0.0015-w  CaCl2  gave  44  drops,  corresponding 
with  the  original  system  and  indicating  that  under  the  conditions  of 
the  experiment  NaCl  and  CaCl2  exert  an  antagonistic  or  compensa- 
tory effect  upon  one  another  in  the  molecular  ratios  of  100  :  1. 

In  similar  experiments  with  other  electrolytes,  anesthetics,  etc.,  the 
ratios  in  which  antagonistic  effects  were  produced  corresponded 
sufficiently  with  those  in  which  the  substances  in  question  exerted 
antagonistic  effects  on  marine  and  other  organisms  as  to  suggest  the 
possibility  that  these  physical  systems  may  afford  a  crude  model  of 
the  mechanism  underlying  the  control  of  permeability  in  protoplasm. 
Salts  of  magnesium  and  other  substances  which  exhibit  abnormalities 


40  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

in  biological  systems  exerting  under  varying  conditions  a  protective 
or  destructive  effect  on  the  protoplasmic  film  exhibit  similar  abnor- 
malities in  emulsion  and  drop  systems.  Magnesium  salts  function 
as  protective  agents  hke  calcium  salts  when  added  to  a  soap  solution 
which  is  passed  through  oil;  but  as  destructive  agents  like  NaOH, 
NaCl  or  KCl  when  added  to  a  dilute  solution  of  NaOH  which  is 
passed  through  an  oil  containing  fatty  acid. 

Closely  parallel  results  were  observed  between  the  drop  system  de- 
scribed above,  the  process  of  blood  coagulation,  the  lethal  dose  of 
given  electrolytes  in  mice  when  injected  intravenously,  the  hemolysis 
of  blood  corpuscles  by  complement  and  amboceptor,  etc.,  a  common 
critical  point  being  observed  in  these  widely  diversified  systems. 
Clowes  considers  that  these  experiments  lend  substantial  support  to 
to  the  view  that  while  protoplasm  as  a  whole  consists  of  a  system 
approximating  more  nearly  to  a  dispersion  of  the  non-aqueous  phase 
in  the  aqueous,  the  extreme  marginal  layer  of  protoplasm  may  be 
looked  upon  as  an  emulsion  or  gel-hke  system  consisting  of  two  con- 
tinuous phases  in  which  fluctuations  in  permeability  to  water  and 
water-borne  substances  may  be  caused  by  variations  in  the  propor- 
tions of  metabolic  products,  electrolytes,  etc.,  a  slight  change  in  the 
system  in  the  direction  of  water  surrounded  by  the  non-aqueous  phase 
leading  to  a  diminution  in  permeability,  while  a  change  in  the  reverse 
direction,  towards  a  system  in  which  the  non-aqueous  phase  is  more 
completely  dispersed  in  the  aqueous,  would  lead  to  an  increased 
permeability. 

Substantial  support  is  lent  to  this  point  of  view  by  Osterhout's 
observations  that  the  conductivity  of  Laminaria  tissue  is  raised  by 
exposure  to  a  solution  of  NaCl,  lowered  by  CaCl2,  but  unchanged 
when  exposed  to  a  mixture  containing  100  molecules  of  NaCl  and 
one  of  CaCl2.  Life  can  only  be  maintained  within  certain  ranges  of 
electrical  resistance  or  permeability  and  an  increase  or  decrease 
in  permeability  beyond  given  limiting  points  is  no  longer  reversible 
and  invariably  causes  death.  Wilder  D.  Bancroft:  Jour.  Phys. 
Chem.,  17,  501  (1913).  G.  H.  A.  Clowes:  Proc.  Physiological  Sec- 
tion, International  Medical  Congress,  pp.  105-114,  London,  1913. 
Proc.  Soc.  Exp.  Biology  and  Medicine,  11,  pp.  1-3,  4-5,  6-8,  8-10 
(1913).  Jour.  Physical  Chemistry,  20,  p.  407  (1916).  Proc.  Soc. 
Exp.  Biology  and  Medicine,  13,  pp.  114-118  (1916).  Science,  43, 
pp.  750-757  (1916).     Tr.] 


CHAPTER  HL 

SIZE  OF  PARTICLES,  MOLECULAR  WEIGHT,  OSMOTIC  PRESSURE, 

CONDUCTIVITY. 

For  the  chemist  wishing  to  discover  the  constitution  of  a  chemical 
substance,  the  determination  of  the  molecular  weight  is  of  great  im- 
portance. Much  time  has  thus  been  spent  on  determining  the 
molecular  weights  of  the  bio-colloids,  such  as  albumin,  starch, 
hemoglobin,  etc.,  and  in  the  following  pages  we  shall  show  what 
probability  for  success  attends  these  efforts. 

A  soluble  substance,  placed  in  a  suitable  medium  which  produces 
no  chemical  change,  distributes  itself  uniformly.  In  the  case  of 
crystalloids,  it  is  impossible  by  either  optical  or  mechanical  means 
to  recognize  the  particles  into  which  it  splits  up.^  We  shall  see  that 
crystalloids  are  frequently  broken  up  into  their  molecules.  Many 
colloids  are  soluble  also.  If  we  examine  their  solutions  in  the  ultra- 
microscope  which  permits  a  hundred  thousand  fold  magnification,^ 
we  can  recognize  numerous  particles.  In  the  case  of  artificial  colloids 
(gold  and  silver  hydrosols)  in  which  we  are  certain  that  all  the  dis- 
solved particles  are  visible,  according  to  R.  Zsigmondy  we  are  in  a  pos- 
ition, as  will  be  shown  by  the  following  considerations,  to  calculate 
the  approximate  weight  of  each  individual  particle.  Let  1  gm.  of 
colloid  be  dissolved  in  1  liter  of  water,  then  every  1  cubic  millimeter 
contains  1/1000  milligram  of  colloid.  If  by  counting  under  the  ultra- 
microscope,  we  determine  that  each  cubic  millimeter  contains  1000 
particles,  we  know  that  each  particle  weighs  one  millionth  of  a  mil- 
ligram. We  can  easily  calculate  the  diameter  of  a  particle  by 
supplying  the  specific  gravity  and  assuming  that  each  particle  is  a 
sphere.  However,  as  soon  as  we  become  uncertain  whether  all  the 
particles  are  visible,  which  is  the  case  with  most  hio-colloids,  the  opti- 
cal method  fails  us.  Under  these  circumstances,  we  can  determine 
the  size  of  the  particles  by  ultrafiltration.  If  we  sift  grains,  we  know 
that  those  which  pass  through  are  smaller,  and  those  which  are 

1  [Optical  inhomogeneity  of  sugar  solution  has  been  demonstrated.  Van  Calcar 
and  L.  de  Bruyn  separated  sodium  sulphate  from  solution  by  high  speed  centrif- 
ugation.     Tr.] 

2  [The  ultramicroscope  makes  visible  otherwise  invisible  particles  but  does  not 
magnify  beyond  the  power  of  its  component  compound  microscope.     Tr.] 

41 


42  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

held  back  are  larger  than  the  meshes  of  the  sieve.  If  we  know  the 
size  of  the  meshes  and  have  several  sieves  with  meshes  of  different 
size,  it  is  easy  to  determine  the  average  size  of  the  grains  by  letting 
them  pass  through  the  different  sieves.  H.  Bechhold's  determina- 
tion of  the  size  of  the  particles  depends  on  this  principle.  Ultrafilters 
(jelly  filters)  with  different  sized  pores  serve  as  sieves  (see  pp.  99  et  seq.). 
Since  there  are  several  methods  for  measuring  the  size  of  the  pores 
(see  p.  100),  it  is  possible  to  determine  definite  limits  for  the  size  of 
the  colloid  particles.^ 

Are  the  particles  thus  found  identical  with  molecules? 

In  the  case  of  metal  sols  we  can  immediately  say,  no.  We  know 
the  molecular  weight  of  metals  and  understand  from  it  that  there  is 
no  prospect  of  directly  seeing  the  molecules  of  the  elements  with 
our  present  instruments.  According  to  E.  Riecke  gold  particles 
of  1  iJifj.  diameter  have  a  molecular  weight  of  300,000,  but  the  molec- 
ular weight  of  gold  is  probably  only  197,  and  the  smallest  particles 
we  can  see  have  a  diameter  of  5  ijljjl.  It  follows,  therefore,  that 
every  recognizable  ultramicroscopic  particle  consists  of  thousands  of 
molecules. 

What  are  the  facts  in  the  case  of  particles  whose  size  is  determinable 
hy  ultrafiltration?  Since  albumin,  starch,  etc.,  have  unusually  large 
molecules,  it  is  probable  that  in  them  the  molecule  and  particle  size, 
as  determined  by  ultrafiltration,  are  identical.  This  is  all  the  more 
likely  since  these  bio-colloids,  like  crystalloids,  are  distributed  by 
means  of  the  action  of  the  solvent,  whereas  the  metal  hydrosols  are 
brought  into  such  minute  divisions  only  by  artificial  means. 

But  what  is  a  molecule?  It  is  the  smallest  portion  of  a  compound 
or  of  an  element  that  may  exist  alone.  If  we  split  a  molecule  of 
common  salt  we  no  longer  have  a  molecule  of  NaCl  but  an  atom 
of  Na  and  an  atom  of  CI.  If  we  divide  an  albumin  molecule,  we 
still  have  complicated  atom  complexes  but  we  have  albumin  no 
longer.  Molecular  weight  is  the  weight  of  a  molecule  compared  to 
that  of  an  atom  of  hydrogen  which  equals  unity.  Consequently  we 
are  measuring  not  absolute,  but  relative  sizes.  The  molecular 
weight  is  determined  by  purely  chemical  means.  If,  for  example, 
we  find  in  sodium  benzoate,  that  there  are  7  atoms  of  carbon 
(7  X  12  =  84),  5  atoms  of  hydrogen  (5  X  1  =  5),  2  atoms  of  oxygen 
(2  X  16  =  32)  and  one  atom  of  sodium  (1  X  23  =  23),  we  should 
know  that  the  molecular  weight  must  be  at  least  144,  because  half 
atoms  do  not  exist.  The  molecular  weight  might  in  fact  be  two 
or  three  times  as  large,  which  would  have  to  be  determined  by  other 

1  [J.  Alexander  has  recently  proposed  measurement  of  particle  size  by  high 
speed  centrifugation,  "  ultracentrifugation."    Tr.] 


SIZE  OF  PARTICLES  43 

chemical  investigations  and  determinations  on  other  chemical 
compounds  of  benzoic  acid. 

From  similar  considerations  the  minimum  value  for  the  molecular 
weight  of  certain  albumins  are  obtained.  If  a  protein  contains  one 
per  cent  of  sulphur,  then  its  molecular  weight  must  be  3200  times 
heavier  than  that  of  hydrogen.  (The  atomic  weight  of  S  =  32.) 
But  there  is  every  reason  to  believe  that  there  are  at  least  two 
atoms  of  sulphur  in  egg  albumen,  because  one-half  of  the  sulphur  is 
easily  split  off,  whereas  the  other  splits  off  with  difficulty.  Thus  egg 
albumen,  with  one  and  three-tenths  per  cent  sulphur  shows  a  molec- 
ular weight  of  4900,  and  oxyhemoglobin  a  molecular  weight  of 
14,800.  Oxyhemoglobin  contains  0.4  to  0.5  per  cent  of  iron;  pro- 
vided it  contains  but  one  atom  of  iron,  its  molecular  weight  mufet  be 
11,200  to  14,000.  The  figures  obtained  approach  one  another  very 
closely. 

Another  method  of  obtaining  the  molecular  weight  is  based  on 
Avogadro's  law.  This  law  says:  "At  the  same  temperature  and 
equal  pressure,  different  gases  contain  the  same  number  of  molecules 
per  liter."  Thus  from  the  weight  of  a  gas  or  of  a  vaporized  sub- 
stance, the  molecular  weight  can  be  determined,  if  we  compare  its 
weight  with  that  of  an  equal  volume  of  hydrogen  gas.  Avogadro's 
law  was  generalized  by  J.  H.  van't  Hoff  and  extended  to  solutions. 
According  to  this  generalization  the  "osmotic  pressure"  of  a  dis- 
solved substance  is  proportionate  to  the  number  of  the  dissolved 
molecules  and  is  as  large  as  if  the  substance  were  vaporized.  If  a 
sugar  solution  is  placed  in  a  porous  clay  cell  which  is  so  dense  that 
water  but  not  sugar  may  pass  in  and  out,^  the  sugar  tries  to  expand 
like  a  gas  and,  as  a  result,  water  enters  the  cell  and  the  solution  rises 
in  it.  If  a  vertical  tube  has  been  attached  to  the  cell,  the  osmotic 
pressure  of  the  solution  may  be  measured  directly  from  the  height  to 
which  the  fluid  rises.  There  are  indirect  methods  also,  the  under- 
lying principles  of  which  we  cannot  discuss  here.  They  depend  on 
the  fact  that  in  proportion  to  the  osmotic  pressure  the  boiling  point 
is  raised  and  the  freezing  point  lowered.  In  ideal  cases  these  changes 
are  strictly  proportionate  to  the  concentration  of  the  dissolved  sub- 
stance in  just  the  same  way  that  the  original  volume  of  an  ideal 
gas  is  reduced  to  one-half  by  double  the  pressure  and  to  one-third 
by  three  times  the  pressure.  Consequently  by  determining  the 
freezing  or  boiling  point  of  a  solution,  molecular  weight  may  be  de- 
termined. In  the  case  of  crystalloids  this  method  is  preferred  to  the 
direct  reading  of  the  osmotic  pressure  for  the  following  reasons:  It 
is  exceedingly  difficult  to  prepare  a  cell  that  really  holds  back  crystal- 
^  Such  a  chamber  is  said  to  be  semipermeable. 


44  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

loids,  and,  on  this  account,  the  danger  of  considerable  error  is  always 
present.  Moreover,  in  solutions  with  ordinary  molecular  weights,  the 
osmotic  pressures  are  so  great  that  very  bulky  apparatus  would  be 
required.  Thus,  for  example,  the  osmotic  pressure  of  a  1  per  cent 
aqueous  solution  of  sugar  at  15.5°  C.  is  actually  0.685  atmosphere. 

The  difficulties  in  the  direct  measurement  of  osmotic  pressure  of 
crystalloids  do  not  exist  however,  in  the  case  of  colloids.  Almost 
any  membrane  keeps  back  colloids  and  the  small  rises  are  easily 
measured.  In  order  to  remove  the  sources  of  error  due  to  the  pos- 
sible presence  of  crystalloids,  we  employ  membranes  which  are  per- 
meable for  crystalloids  instead  of  semipermeable  ones  (collodion 
sacs,  animal  membranes). 

These  physical  methods  for  determining  the  molecular  weight  rest 
on  the  assumption  that  the  substance  in  solution  is  really  broken  up 
into  molecules  (colloids  cannot  be  vaporized).  This  condition  does 
not  always  exist  in  the  case  of  crystalloids  and  only  exceptionally 
with  colloids.  With  crystalloids  these  methods  yield  figures  that 
are  either  too  low  or  too  high.  The  former  will  occur  if  the  sub- 
stance is  incompletely  split  up  —  if  two,  three  or  more  molecules 
continue  to  be  united  in  solution.  Under  these  circumstances  we  ob- 
tain one-half,  one-third,  etc.,  the  osmotic  pressure  that  a  molecular 
subdivision  would  show.  The  ultramicroscope  and  ultrafiltration 
reveal  in  many  solutions  of  bio-colloids  particles  of  such  a  size  as 
show  no  molecular  subdivision  by  other  methods;  we  may  assume 
therefore  that  colloidal  solutions  for  the  most  part  contain  molecular 
groups,  and  that  there  is  no  prospect  of  determining  the  true  molec- 
ular weight  by  osmotic  methods. 

The  osmotic  method  yields  a  deceptively  low  molecular  weight 
if,  for  example,  the  substance  is  dissociated  further  than  into  mole- 
cules. This  occurs  in  the  case  of  electrolytes.  A  very  dilute  NaCl 
solution  that  has  dissociated  into  Na  and  CI  ions  shows  twice  the 
true  osmotic  pressure,  so  that  the  molecular  weight  might  be  set  at 
half  its  real  value.  In  this  respect  we  may  also  make  mistakes  with 
colloids  whenever  they  are  ionized.  The  osmotic  method  shows  us  only 
into  how  many  fragments  a  molecular  complex  breaks  up  in  the  par- 
ticular solution.  It  may  give  either  minimum  or  maximum  values  for 
the  molecular  weight.  Even  in  the  case  of  crystalloids,  the  method 
must  be  employed  with  due  consideration  of  all  the  conditions  in- 
volved; for  colloids  it  may  be  exceedingly  deceptive.  We  know  at 
the  outset,  because  of  the  enormous  molecular  weight  of  colloids,  that 
the  lowering  of  the  freezing  point  and  the  elevation  of  the  boiling 
point  must  be  very  small  indeed,  requiring  most  delicate  measure- 
ments.    The  matter  becomes  still  further  complicated  by  the  fact 


SIZE  OF  PARTICLES 


45 


that  crystalloids  are  adsorbed  by  colloids  and  cannot  be  completely 
removed  by  dialysis.  Each  crystalloid  molecule  or  each  crystalloid 
ion  may  thus  falsely  represent  the  osmotic  pressure  of  a  colloid  mole- 
cule having  perhaps  a  thousand  times  its  mass. 

The  coefficient  of  diffusion  may  be  employed  in  the  determination 
of  the  molecular  weight  of  crystalloids,  but  in  the  case  of  colloids 
it  gives  information  concerning  only  the  average  size  of  the  particles. 
The  method  is  not  much  impaired  by  the  increase  in  the  size  of  the 
molecule,  because  it  is  only  the  square  of  the  coefficient  of  diffusion 
which  diminishes  proportionately  to  this  increase.  The  adsorption 
of  crystalloids,  on  the  contrary,  is  also  in  this  case  a  source  of  error 
because  every  crystalloid  molecule  or  ion  acts  as  a  team-mate  of  its 
colloid  particle  and  hastens  its  rate  of  diffusion.  The  objection  to 
the  principles  governing  the  calculation  are  mentioned  on  page  53. 

Before  we  come  to  concrete  examples  we  shall  mention  one  other 
method  which  may  enlighten  us  concerning  the  particle  content  of 
a  solution  —  the  conductivity.  In  a  solution  the  electric  current  is 
carried  only  by  the  electrically  charged  particles  (ions).  In  an  NaCl 
solution  this  is  done  by  the  Na  and  CI  ions;  in  a  Na2S04  solution, 
2  Na  ions  and  1  SO4  ion,  that  is  3  ions,  take  part.  The  assimiption 
is  that  many  molecules  are  completely  or  almost  completely  split  into 
ions;  this  actually  occurs  in  the  case  of  strong  electrolytes  when  in 
great  dilution.  The  conductivity  thus  affords  us  fractions  of  the 
molecular  weight:  minimum  figures  (values  less  than  actual). 

My  chief  purpose  in  making  these  statements  is  to  show  what 
facts  may  be  deduced  from  the  various  methods  used  in  determining 
the  molecular  weight;  they  give  only  limiting  values,  so  that  no  con- 
clusion is  to  be  drawn  from  any  one  method. 

The  following  remarks  will  show  what  difficulties  stand  in  the 
way  when  we  try  to  learn  the  size  of  the  colloid  molecule. 

Among  the  colloids  whose  chemical  composition  is  best  known 
are  the  soaps. 

As  was  found  by  F.  Krafft  and  A.  Smits,  very  dilute  soap  solu- 
tions showed  a  well-marked  rise  in  boiling  point;  but  this  did  not 
rise  in  proportion  to  the  concentration  of  the  soap,  as  may  be  seen 
from  the  following  table  by  A.  Smits  for  sodium  palmitate: 


Concentration  in  mols. 

Rise  in  boiling  point, 
°C. 

0.0282 
0.1128 
0.2941 
0.5721 

0.024 
0.045 
0.050 
0.060 

46  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Though  the  concentration  is  increased  twenty  fold,  the  boihng 
point  rises  only  two  and  one-half  times.  In  a  solution  of  19,5  per 
cent  sodium  stearate,  F.  Krafft  found  absolutely  no  rise  in  the 
boiling  point  as  compared  to  pure  water. 

Let  us  examine  the  conclusion  of  W.  Biltz  and  A.  v.  Vegesack 
based  on  their  critical  study  of  the  osmotic  method.  True  colloids, 
like  iron  oxid  and  tungstic  acid  show  a  small  osmotic  pressure,  only 
so  long  as  they  contain  electrolytes.  As  the  electrolyte  vanishes, 
the  colloid  particles  aggregate  to  larger  complexes  which  then  cease 
to  show  any  osmotic  pressure.  For  the  existence  of  these  colloids, 
some  electrolyte  content  is  an  absolute  necessity. 

When  these  investigators  studied ''  colloid  electrolytes,"  particularly 
colloid  color  salts  (congo  red,  night  blue  and  benzopurpurin)  whose 
constitution,  molecular  weight,  etc.,  were  determined  by  chemical 
methods,  they  obtained  results  which  especially  in  the  case  of  congo 
red  must  be  closely  examined.  Congo  red  has  the  formula  C32H22N6- 
S206Na2,  and  being  a  sodium  disulphonate,  is  a  strong  electrolyte. 
Its  molecular  weight  (M)  =  696.  On  account  of  its  electrolytic  dis- 
sociation into  3  ions  (2  crystalloid  and  1  colloid)  we  would  expect 
an  osmotic  pressure  three  times  as  much  as  its  molecular  weight 
would  indicate.  Instead  of  this  W.  M.  Bayliss  and  also  W.  Biltz 
and  A.  v.  Vegesack  as  well  as  Donnan  and  Harris  obtained  by 
dialysis  against  pure  water  a  pressure  which  was  approximately  5  per 
cent  lower  than  would  have  been  obtained  had  the  undissociated 
molecule  been  active.  The  explanation  is  not  difficult.  Let  us  desig- 
nate by  R,  the  acidic  color  radical  of  congo  red,  then  congo  red  has  the 
formula  Il.Na2.  In  solution  a  portion  becomes  ionized  into  R  and 
Na  Na,  of  which  some,  even  though  possibly  only  a  small  fraction 
forms  with  the  H  and  OH  ions  of  the  water  RH  (color  acid)  and 
NaOH.  This  occurrence  would  be  without  much  influence  in  chang- 
ing osmotic  pressure  if  the  measurement  was  made  in  a  closed  vessel 
in  which  the  equilibrium  was  undisturbed.  As  a  matter  of  fact  the 
measurement  is  made  in  a  membrane  permeable  for  electrolytes. 
Consequently  the  NaOH  which  has  been  formed  diffuses  away  and 
some  fresh  color  acid  (RH)  may  be  formed.  This  process  continues 
until  practically  only  color  acid  remains  in  the  membrane.  Con- 
sequently in  this  instance  we  have  not  measured  the  high  osmotic 
pressure  of  the  electrolytically  strongly  dissociated  color  salt  but  that 
of  the  practically  undissociated  color  acid.  If  measured  against  outer 
water  containing  electrolytes,  it  yields  a  very  much  lower  osmotic 
pressure,  equivalent  to  a  value  for  M  of  2088.  We  shall  thoroughly 
understand  this  occurrence  when  we  have  become  more  familiar  with 
the  equilibria  of  membranes.     See  page  59. 


SIZE  OF  PARTICLES  47 

We  shall  only  indicate  here,  that  when  the  colloid  electrolytes 
(thus  BiLTZ  designates  salts  in  which  one  ion  is  a  colloid)  and  the 
electrolyte  in  the  outer  water  have  originally  the  same  osmotic 
pressure,  there  is  a  gradual  penetration  of  the  outer  electrolytes  but 
the  colloid  electrolytes  cannot  escape.  Consequently  the  osmotic 
pressure  in  the  cell  is  the  resultant  of  osmotic  pressure  of  the  colloid 
electrolytes  plus  that  of  the  electrolytes  which  have  entered.  If  the 
latter  have  an  ion  in  common  with  the  colloid  electrolytes,  we  do  not 
find,  as  might  have  been  expected,  that  there  is  an  equal  division  of 
the  true  electrolytes  {e.g.,  NaCl),  but  on  the  contrary,  outside  the 
osmometer  there  is  proportionately  more  NaCl  the  more  dilute  the 
NaCl  solution  is  (see  page  62) .  The  osmotic  pressure  of  the  colloid 
electrolytes  is  consequently  depressed. 

From  this  it  follows  that  the  values  determined  by  the  direct 
osmotic  methods  require  revision.  E.  H.  Starling  obtained  4  mm, 
Hg  osmotic  pressure  for  serum  colloid  in  1  per  cent  serum,  thus  ex- 
pressing an  apparent  molecular  weight  of  about  50,000.  E.  W.  Reid 
obtained  a  pressure  of  369  mm.  Hg  for  a  1  per  cent  hemoglobin  solu- 
tion from  which  is  deduced  an  apparent  molecular  weight  of  about 
65,000,  a  figure  which  approaches  the  values  obtained  by  the  diffusion 
method  by  R.  0.  Herzog  and  Sv.  Arrhenius. 

These  figures  are  4  to  10  times  greater  than  those  determined  for 
the  molecular  weight  by  chemical  means. 

As  a  matter  of  fact  the  theory  of  the  direct  measurement  of  osmotic 
pressiu-e  is  so  difiiciilt  that  I  do  not  know  any  results  which  are  not  sus- 
ceptible of  adverse  criticism  (see  F.  G.  Donnan's  theory).  As  the  re- 
sult of  direct  measurement,  we  know  that  colloid  solutions  actually  have 
an  osmotic  pressure  and  that  it  increases  with  the  amount  of  dispersion. 
Theoretical  considerations,  however,  show  us  that  this  osmotic  pressure 
must  be  low.  Theoretically,  all  solutions  which  contain  the  same 
number  of  particles  of  the  dissolved  substance  exert  the  same  osmotic 
pressure.  Thus  all  normal  solutions  (leaving  out  of  consideration 
dissociations,  associations  and  other  changes)  exert  the  same  osmotic 
pressure,  namely,  22.4  atmospheres.  Normal  solutions  are  such  as 
contain  the  same  number  of  molecules,  namely,  one  gram  molecule 
per  liter.  A  normal  salt  solution  is  one  containing  58.5  gm.  NaCl 
per  liter  and  a  normal  hydrogen  solution  is  one  containing  2  gm.  of 
hydrogen  per  liter  (theoretically).  By  various  means  which  yield 
rather  concordant  results  it  has  been  determined  that  2  gm.  H  contain 
6.1  X  10"^  molecules^  or  fragments  of  molecules  or  molecular  com- 
plexes, i.e.,  particles  in  solution  exerting  22.4  atmospheres  of  osmotic 

^  This  figure  (6.1  X  10-')  is  called  Avogadro's  figure,  and  the  various  methods 
for  deriving  it  give  quite  uniform  values. 


48  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

pressure.  When  we  consider  that  the  finest  gold  particles  of  a  col- 
loidal gold  solution  have  a  diameter  of  only  2/j,fjL^  (a  dimension  one- 
fifth  that  of  ultramicroscopic  visibility);  according  to  Svedberg  in 
order  to  make  a  normal  colloidal  gold  solution  we  should  have  to  cram 
50  kilos  of  gold  into  a  liter,  which  would  have  to  contain  6.1  X  10^^ 
such  gold  particles.  It  is  naturally  impossible  to  do  this;  the  most 
that  can  be  dissolved,  experimentally,  is  one  gm.  per  liter.  Under 
favorable  conditions  such  solutions  have  an  osmotic  pressure  of 
4.5  X  10~*  atmosphere,  i.e.,  they  would  rise  4.65  mm.  in  an  osmom- 
eter, or  be  in  equilibrium  with  electrolytes  which  modify  the  electro- 
lytic or  hydrolytic  dissociation  of  colloid  electrolytes. 

It  is  unnecessary  to  catalogue  all  the  dozens  of  fruitless  investi- 
gations of  the  molecular  weight  of  colloids  by  physical  methods. 
They  either  gave  surprisingly  low  molecular  weights,  when  it  could 
be  shown  on  testing  that  the  colloid  was  contaminated  with  crystal- 
loids, or  the  values  were  so  small  (the  molecular  weight  so  large) 
that  they  fell  within  the  limits  of  error  of  observation,  which  means 
that  it  became  doubtful  whether  the  colloid  studied  had  any  osmotic 
pressure  at  all.  The  molecular  weight  is  the  expression  of  a  chemical 
point  of  view  which  cannot  be  determined  by  ^physical  methods  for 
colloids.  What  we  obtain  by  these  methods  are  more  or  less  nu- 
merous groups  of  molecules,  usually  in  adsorption  balance  with  irre- 
movable traces  of  crystalloids  which  cannot  be  separated,  or  which 
are  in  equilibrium  with  electrolytes  that  influence  the  electrolytic 
and  hydrolytic  dissociation  of  the  colloid  electrolytes. 

In  the  present  state  of  the  science  we  may  only  strive  to  determine 
the  size  of  the  particles  in  solutions  of  colloids  having  various  equilibria. 

1  [According  to  Zsigmondy  particles  5mm  in  size  may  be  seen  with  the  aid  of 
strong  sunlight.  —  Tr.] 


CHAPTER  IV. 

PHENOMENA  OF  MOTION. 

Brownian-Zsigmondy  Movement. 

Upon  examining  a  drop  of  milk  under  the  microscope,  we  are  at 
once  struck  by  the  appearance  of  the  fat  droplets  on  account  of  their 
strong  refraction  (a  dark  ring  with  a  brilliant  center).  It  is  seen 
that  they  exhibit  a  certain  oscillation  (trembling).  This  chu,racter- 
istic  oscillating  movement  is  more  intense  with  the  smaller  droplets 
(Fig.  5),  whereas  those  having  a  diameter  of  more  than  4  (j.  do  not 
show  it  at  all.     The  phenomenon  is  named  after  the  English  botan- 


FiG.  5.     Brownian  movement  of  milk  globules.     (From  O.  Lehman.) 

ist,  Robert  Brown,  who  discovered  it  as  early  as  1827,  in  an  aqueous 
suspension  of  plant  pollen.  It  may  be  observed  in  every  suspension 
or  emulsion  which  is  sufficiently  fine.  Particles  of  1  ju  diameter  show 
a  radius  of  translation  of  1  ^t.  The  ''dance  of  the  motes,"  the  rush- 
ing hither  and  thither  of  the  bright  particles  observed  in  the  ultra- 
microscope,^  is  nothing  else  than  an  enormously  exaggerated  Brownian 
movement,  due  to  the  fact  that  the  particles  are  much  smaller  than 
those  that  may  be  seen  under  the  microscope.  Particles  of  10  to  50  /jlh 
have  a  speed  of  more  than  100  /x  per  second.      These  movements 

^  R.  Lorenz  correctly  calls  attention  to  the  fact  that  the  great  advance  in  our 
science  does  not  date  from  Brown  who  observed  the  "oscillations"  of  microscopic 
particles  but  from  R.  Zsigmondy  who  recognized  that  particles  of  molecular 
dimensions  are  in  a  similar  mobile  state.     (Frankforter  Ztg.  4.6.11,  I  Morgenbl.) 

49 


50  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

seen  under  the  ultramicroscope  are  comparable  to  the  dance  of  the 
molecules  in  accordance  with  the  Kinetic  Theory  of  Gases. 

The  speed  of  the  particles  is  dependent  on  the  viscidity  of  the 
dispersing  medium  and  increases  with  a  rise  in  temperature.  It  is 
very  pertinent  to  enquire  at  this  point  whether  we  here  see  the  move- 
ments of  the  molecules  themselves.  In  a  certain  sense,  this  may  be 
answered  in  the  affirmative.  Though  we  cannot  yet  say  that  this 
movement  is  inherent  in  the  particles,  i.e.,  that  it  would  be  carried 
out  by  the  particles  themselves,  we  may  assert  that  it  is  caused  by 
blows  from  the  molecules  of  the  solvent. 

A.  Einstein  and  M.  von  Smoluchowski  have  independently  de- 
duced from  the  Kinetic  Theory  of  Gases,  laws  for  the  Brownian 
movement  (extent  of  movement,  influence  of  temperature  and  vis- 
cosity). It  might  be  assumed  a  priori,  that  a  particle  floating  in  a 
fluid  would  remain  at  rest,  for  it  simultaneously  receives  from  all 
sides  an  equal  number  of  impacts  from  molecules.  The  fallacy  of 
this  assumption  is  shown  by  M.  von  Smoluchowski  in  a  very  pretty 
comparison.  If  we  play  roulette  for  a  long  time  the  chances  for 
gaining  and  losing  are  equal  (disregarding  the  banker).  If  we  play 
only  a  short  time  we  win  one  day  and  lose  the  next.  In  other  words 
the  law  of  probabilities  shows  that  the  excess  of  molecular  impacts 
which  reach  a  particle  in  a  given  quarter  suffice  to  give  it  movement 
one  direction  or  another  direction.  The  smaller  the  particle,  the 
greater  is  the  probability  that  the  impacts  will  not  arrest  it  and  the 
stronger  is  its  movement. 

The  formula  of  von  Smoluchowski  as  well  as  that  of  A.  Einstein 

demands  that  -~  be  constant  for  equal  sized  particles; 

A  =  amplitude,     tq  =  viscosity,     T  =  oscillation  time. 

Th.  Svedberg,  by  brilliantly  devised  methods,  measured  these 
values  on  colloid  metals,  in  various  dispersing  media,  and  estab- 
lished the  constants.  It  is  true  that  the  absolute  figures  Ä^the. 
measured  and  for  the  calculated  amplitudes  do  not  exactly  ngree, 
but  they  are  of  the  same  order  of  magnitude;  i.e.,  the  values  found 
are  on  the  average  three  times  as  large  as  those  calculated.  Seddig, 
also,  confirmed  the  quantitative  increase  of  amplitude  accompany- 
ing a  rise  in  temperature. 

This  is  a  remarkable  agreement  between  the  movements  of  small 
particles  seen  with  the  eyes  and  the  hypothesis  of  the  movements 
of  gas  molecules  based  on  scientific  imagination,  which  Kroenig  in 
1856  and  Clausius  in  1857  formulated  mathematically  (kinetic 
theory  of  gases).-    All  investigations  that  have  since  been  undertaken 


PHENOMENA  OF  MOTION  51 

concerning  the  laws  of  gases  and  the  movement  of  colloidal  particles 
have  essentially  agreed  in  showing  that  the  laws  of  gases  proved 
applicable  to  very  dilute  solutions  of  hydrophobe  colloids  and  con- 
versely, that  the  laws  of  gases  could  be  developed  from  the  move- 
ments of  colloid  particles.  Boyle's  law  asserts  that  the  volume  (v) 
of  a  gas  is  inversely  proportional  to  the  pressure  (p)  exerted  upon  it: 
V  :  v'  =  p'  :  p.  According  to  Gay-Lussac's  law  the  volume  change 
of  a  gas  having  the  temperature  (t)  is  v  =  Vo  {I  +  at)  in  which  Vo  is 
the  volume  at  0°  and  a  is  the  coefficient  of  expansion.  From  the 
standpoint  of  molecular  kinetics  under  doubled  pressure  twice  as 
many  moving  particles  are  present  in  the  same  space.  With  increase 
of  temperature  (assuming  the  same  pressure)  at  times  fewer  parti- 
cles are  present  than  in  the  same  gas  volume  at  0°.  This  assumes 
average  values,  though  in  fact,  the  number  of  particles  in  a  definite 
volume  varies  from  m.oment  to  moment.  If  this  assumption  is  correct 
the  average  of  the  "instant  values"  must  give  values  which  sat- 
isfy Boyle-Gay-Lussac's  law.  M.  von  Smoluchowski  developed 
mathemartically  the  relation  between  this  law  and  the  "  instant 
values."  He  obtained  experimental  verification  when  Th.  Svedberg 
counted  the  number  of  particles  for  an  "instant  value"  directly  in 
the  ultramicroscope  and  R.  Lorenz  counted  the  particles  in  cine- 
metagraphs  of  the  ultramicroscopic  field.  The  assumption  also 
bridges  the  gap  between  Thermodynamics,  which  studies  phenomena 
on  the  basis  of  the  involved  energy  and  its  transformations,  and  the 
Kinetic  Molecular  Theory,  which  views  matter  as  the  smallest  possible 
particles  in  motion. 

The  impact  that  our  ultramicroscopically  visible  particles  exert 
against  the  walls  of  a  vessel  is  the  pressure  they  exert,  and  it  is 
measurable  for  a  molecularly  dispersed  system  as  the  osmotic 
pressure. 

The  osmotic  pressure,  a,  function  of  the  mass  and  motion  of  a  sus- 
pension whose  particles  are  visible  and  measurable,  was  shown  by 


^►:mRiN  to  coincide  with  the  requirements  of  the  Kinetic  Theory 
of  Cases  and  of  Thermodynamics;  that  is,  with  its  energy  content  in 
the  form  of  heat. 

The  following  considerations  make  this  clear. .  Under  the  influence  of  gravity 
the  lower  layers  of  the  atmosphere  have  a  greater  density  than  the  upper;  i.e., 
the  number  of  gas  particles  (molecules)  in  1  cc.  is  greater  in  the  immediate 
vicinity  of  the  earth  than  at  higher  altitudes.  This  applies  not  only  for  gases 
but  also  for  solutions  or  suspensions.  " 

J.  Perrin  prepared  a  very  fine  suspension  of  gamboge  which  he  placed  in  a 
tall  cylinder.  Gradually  under  the  influence  of  gravity  an  equilibrium  developed 
in  which  there  was  dense  suspension  at  the  bottom  of  the  cyUnder  with  gradually 
diminishing  concentration  of  particles  in  the  upper  layers  —  an  atmosphere  in 


52 


COLLOIDS  IN  BIOLOGY  AND   MEDICINE 


miniature  (Fig.  6).  By  means  of  the  ultramicroscope  he  counted  particles  in 
each  layer  of  0. 12  mm.  The  (osmotic)  pressure  of  a  particle  may  be  calculated 
by  the  following  formula  applicable  to  the  kinetics  of  gases: 


In 


no 


=  lmgh{l  -l^j; 


r. 


no  and  n  are  the  number  of  particles  counted  in  the  unit  volume  at  the  levels 

0  and  h,  m  =  mass,  g  =  acceleration  due  to 
gravity,  s  =  density  of  particles.  The  value  of  k 
proved  to  be  43  •  lO-i'^. 

If  this  pressure  of  a  single  particle  expresses 
the  pressure  of  a  single  molecule  (in  solution  or  as 

a  gas),  the  equation  k  =  —  appUes. 

Here  N  is  the  number  of  the  molecules  present  in 
a  gram  molecule  as  determined  by  other  methods, 
that  is,  6  •  1023,  T  =  295°,  A;  is  43  •  IQ-^K  From 
this  we  derive  the  value  ß  =  2.1  cal.  instead  of 
1.98  cal.  From  this  the  molecular  weight  of  the 
gamboge  particles  was  proved  to  be  3  •  10'. 

Diffusion. 

If  pure  water  is  layered  over  a  concen- 
trated sugar  or  salt  solution  so  that  there 
is  no  mixing,  it  is  found  that  after  a  certain 
time  (hours  or  days),  the  sugar  or  salt 
passes  into  the  water;  we  say  that  it 
diffuses  into  the  water.  If  one  chooses  a 
colored  salt  solution,  e.gr.,  copper  sulphate, 
the  path  of  the  diffusion  may  be  easily 
observed  by  the  coloration.  It  is  in  prin- 
ciple the  same  process  as  when  one  per- 
mits a  compressed  gas  to  stream  into  the 
air  —  the  distinction  lies  only  in  the  differ- 
ence in  speed. 

It  has  been  shown  that  different  sub- 
stances have  very  different  rates  of  diffu- 
sion which  are  characteristic  for  these 
substances.  These  characteristic  con- 
stants are  called  coefficients  of  diffusion. 
The  coefficient  of  diffusion  expresses  the  amount  of  substance  which 
passes  through  an  area  1  cm.^  per  second  ^  from  a  solution  contain- 
ing 1  part  per  cc. 

^  Because  this  time  is  so  brief  it  is  usually  necessary  to  multiply  the  coefficients  . 
by  a  large  factor  or  to  choose  the  day  as  the  unit  of  time. 


c*.- 


I* 


'  ,.,'f.,. 


Pig.  6.  Mastic  suspension 
showing  the  effect  of  grav- 
ity.    (Perrin.) 


PHENOMENA  OF  MOTION  53 

.J 

It  is  evident  that  these  values  are  free  from  any  hypothetical  con- 
siderations. It  has,  however,  been  shown  that  the  coefficient  of 
diffusion  for  crystalloids  bears  a  certain  relation  to  the  molecular 
weight.  Small  molecules  diffuse  rapidly,  large  ones  slowly.  When 
suitable  formulas  are  used  there  is  very  satisfactory  correspondence, 
provided  the  molecular  weights  are  not  smaller  than  50  nor  larger 
than  500.  A  further  advance  was  made  by  seeking  to  calculate  from 
the  coefficient  of  diffusion  the  molecular  weights  of  colloids  whose  M 
was  unknown.  The  results  were  not  concordant  because  the  moving 
units  in  colloidal  solutions  are  not  "molecules"  but  "particles,"  that 
is,  complexes  of  molecules. 

If  we  connect  the  fact  that  the  coefficient  of  diffusion  decreases 
with  increase  in  the  size  of  the  molecule,  with  what  we  know  about 
the  Browfiian-Zsigmondy  movement,  the  relationship  is  surprising.  We 
have  seen  that  the  movement  is  smaller  as  the  particles  grow  larger 
and  it  is  evident  that,  when  we  layer  water  over  a  metal  hydro- 
sol,  the  strong  translatory  movements  which  we  observe  under  the 
ultramicroscope  must  carry  the  hydrosol  into  the  pure  water.  In 
coarser  suspensions  possessing  only  vibratory  movements,  we  do 
not  expect  diffusion  to  occur.^  Svedberg  measured  the  diffusion 
coefficient  in  different  solutions  of  colloidal  gold  and  calculated  the 
size  of  the  particles  from  the  very  simple  relation  (particle  size  in- 
versely proportional  to  diffusion  coefficient).  The  experiments  were 
carried  out  with  two  gold  solutions  which  contained  particles  of 
1-3mm  and  of  20-30ßiJ,,  directly  measured  ultramicroscopically. 
There  was  a  relatively  good  agreement  between  the  results  as  cal- 
culated and  determined.  A  direct  relationship  between  the  Brown- 
ian-Zsigmondy  movement  and  the  coefficient  of  diffusion  cannot  be 
experimentally  established  by  methods  free  from  criticism,  because 
the  hydrophobe  hydrosols  (e.g.,  colloidal  gold,  platinum  and  the  like) 
which  may  be  counted  under  the  ultramicroscope  cannot  be  prepared 
entirely  free  of  crystalloids.  Since  every  crystalloid  molecule  which 
is  attached  to  a  colloid  particle  must  greatly  accelerate  the  diffusion 
of  the  latter,  we  are  confronted  with  a  source  of  error  that  is  uncon- 
trollable. 

A  series  of  coefficients  of  diffusion  have  also  been  measured  for 
hydrophile  colloids,  which  though  they  have  not  the  exactitude 
possessed  by  those  of  crystalloids,  reveal  a  remarkable  constancy 
so  that  they  may  be  considered  characteristic  for  the  substance 
under  consideration  (Sv.  Arrhenius,  R.  O.  Herzog,  Euler, 
Öholm)  . 

^  For  the  mathematical  relations  between  diffusion  coefficient,  molecular 
weight  and  molecule  or  particle  diameter  see  R.  O.  Herzog  and  L.  W.  Oholm. 


54  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

This  may  be  illustrated  by  the  following  figures: 


Diffusion 
coefficient 
D  at  20°  C. 

Molecular  weight 

Particle 

diameter 

in  MM. 

Substance. 

otherwise 
determined. 

calculated 
from  D. 

Observer. 

Urea 

110 
73 

66 

38 

14 

10.5 
7 
7 

6.6 
5.9 
3.6 
3.3 

60 

92 

110 

342 

j    973? 
I  2612? 

40 

91 

111 

337 

2,430 

4,440 
10,000 
10,000 
11,200 
14,200 
37,700 
44,900 

0.34 
0.51 
0.57 
0.98 

2.65 

3.57 
5.36 
4.60 

4.88 
5.46 
8.88 
9.76 

Öholm 

Glycerin 

Resorcin 

a 

Cane  sugar 

Graham- 

Inulin 

Stephan 
Öholm 

Dextrin 

Soluble  starch 

(( 

Pepsin 

Rennin 

Egg  albumen 

Emulsin 

Herzog 

Invertin 

a 

All  the  facts  mentioned  above  indicate  what  Einstein  emphasized 
even  in  1905,  that  there  is  a  gradual  transition  from  crystalloid  to 
colloid  solutions,  and  that  the  translatory  movements  of  the  particles 
of  a  colloid  correspond  to  the  diffusion  of  crystalloid  molecules. 

[Sir  William  Ramsay,  in  a  paper  entitled  "  Pedetic  Motion  in 
Relation  to  Colloidal  Solutions"  published  in  "Chemical  News," 
Vol.  65,  p.  90  [1892],  stated  as  follows: 

"  I  am  disposed  to  conclude  that  solution  is  nothing  but  subdivi- 
sion and  admixture,  owing  to  attractions  between  solvent  and  dis- 
solved substance  accompanied  by  pedetic  motion;  that  the  true 
osmotic  pressure  has,  probably,  never  been  measured;  and  that  a 
continuous  passage  can  be  traced  between  visible  particles  in  suspen- 
sion, and  matter  in  solution;  that,  in  the  words  of  the  old  adage, 
Natura  nihil  fit  per  saltum."     Tr.] 

Diffusion  in  Jellies. 

Hitherto  we  have  considered  only  diffusion  in  pure  aqueous  solu- 
tion; in  the  organism,  however,  it  occurs  in  a  more  or  less  dense 
colloidal  medium.  When  the  concentration  of  the  colloid  is  not 
very  great,  the  diffusion  is  not  much  impeded.  Until  recently  it 
was  even  believed  that  diffusion  of  a  crystalloid  solution  in  a  jelly, 
e.g.,  gelatin  or  agar,  occurred  just  as  rapidly  as  in  pure  water.  This 
was  the  result  of  employing  unsuitable  experimental  methods. 
The  investigations  of  H.  Bechhold  and  J.  Ziegler,*^  Kurt  Meyer,* 
Peter  Nell*  and  L.  W.  Öholm  showed  definitely  that  electrolytes 
and  non-electrolytes  experience  a  resistance  in  jellies  which  reduced 
the  rate  of  diffusion  by  obstructing  their  paths,  and  that  the  inter- 
ference increased  if  the  gel  became  more  concentrated. 


PHENOMENA   OF  MOTION  55 

Even  the  age  of  the  jelly  may  have  an  influence.  Thus,  F.  Stof- 
fel *  showed  (from  H.  Zangger's  laboratory)  that  the  diffusion- 
path  of  crystalloids  in  gelatin  which  was  rapidly  solidified  is  greater 
than  it  is  in  gelatin  which  was  slowly  solidified,  but  that  this  becomes 
equalized  after  several  days. 

The  rate  of  diffusion  may  be  delayed  or  hastened  through  the 
presence  of  a  third  substance.  This  affects  the  diffusion  in  jellies  to 
a  much  greater  extent  than  in  liquids.  On  page  69  et  seq.,  we  shall 
see  that  chlorin,  iodin,  nitrate,  and  other  ions,  urea,  etc.,  favor  swell- 
ing; on  the  other  hand,  sulphate,  citrate,  and  other  ions  as  well  as 
alcohol,  sugar,  etc.,  as  compared  with  pure  water,  diminish  swelling, 
so  that  to  a  certain  extent  the  meshes  of  the  colloid  network  may 
be  opened  or  closed.  It  is  easy  to  understand  that  diffusion  will 
occur  less  rapidly  through  narrow  meshes  than  through  wide  ones. 
That  such  an  influence  on  diffusion  actually  occurs  was  experimen- 
tally shown  by  H.  Bechhold  and  J.  Ziegler.  They  showed  that  the 
permeability  of  gelatin  and  agar  jellies  for  electrolytes  and  non- 
electrolytes  was  increased  by  urea,  whereas  it  was  diminished  by 
sodium  sulphate,  grape  sugar,  glycerin  and  alcohol.  An  increase  is 
also  produced  by  sulpho-groups,  according  to  Böhi.* 

It  is  evident  that  every  substance  which  increases  the  permeability 
of  other  substances,  paves  the  way  for  its  own  passage  as  weh.  If 
a  jelly  has  been  saturated  with  urea,  the  later  coming  particles  of 
urea  will  diffuse  more  rapidly;  conversely,  sodium  sulphate  and 
grape  sugar  particles  obstruct  by  their  influence  on  the  gel  the  pas- 
sage of  the  subsequent  particles. 

There  are  other  ways  in  which  diffusion  in  a  gel  may  be  dis- 
tinguished from  that  in  aqueous  solution.  We  know  from  Chapter  II 
that  colloids  adsorb  other  substances  to  a  greater  or  less  extent. 
By  this  means  diffusion  may  be  more  or  less  impeded  and  under 
certain  circumstances  even  entirely  arrested.  This  may  be  ob- 
served with  ease  in  the  diffusion  of  dyes.  H.  Bechhold  and  J. 
Ziegler*^  showed  that  gelatin  was  deeply  stained  with  methylene 
blue  and  thus  the  diffusion  in  the  gelatin  was  impeded;* whereas  the 
juice  of  red  beets  is  a  dye  which  is  not  noticeably  adsorbed.  Finally, 
if  we  observe  that  adsorption  is  strongly  influenced  by  the  presence 
of  salts  and  non-electrolytes,  and  that  an  effect  on  diffusion  is  thus 
exerted,  we  shall  see  what  great  complications  may  appear  when 
diffusion  occurs  in  colloid  media. 

Though  a  diffusion  of  colloids  in  aqueous  media  was  long  doubted 
the  diffusion  of  colloids  in  jellies  was  positively  denied.  Thomas 
Graham  held  it  to  be  characteristic  of  colloids  that  they  were  ar- 
rested by  other  colloids.  H.  Bechhold*^  called  attention  to  the 
*  [Graham  recognized  the  slow  diffusion  of  colloids.     Tr.] 


56  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

ability  of  true  albumins  to  diffuse  into  gelatin  jellies.  This  fact  was 
demonstrated  by  means  of  the  precipitin  reaction.  If  goat  serum 
is  mixed  with  rabbit  serum  nothing  noteworthy  occurs.  If  the 
rabbit  has  previously  been  injected  with  goat  serum  and  the  serum 
of  the  previously  treated  rabbit  (called  ''goat-rabbit  serum")  is 
mixed  with  goat  serum  there  occurs  a  precipitation  of  an  albumi- 
nous substance,  called  ''precipitin."  Bechhold  mixed  a  1  per  cent 
gelatin  solution  containing  0.85  per  cent  NaCl  with  an  equal  vol- 
ume of  goat-rabbit  serum.  The  jelly  was  sohdified  in  the  ice  box 
and  goat  serum  was  layered  over  it.  At  the  end  of  24  hours  a  cloudy 
precipitate  formed  in  the  gelatin  which  in  the  course  of  120  hours 
penetrated  as  far  as  5  mm.  The  same  phenomena  occurred  when 
the  gelatin  was  mixed  with  goat  serum  and  goat-rabbit  serum  was 
layered  over  it.  Thus  in  both  cases  actual  constituents  of  the  serum 
had  diffused  into  the  gelatin. 

Similarly,  Sv.  Arrhenius*  and  Th.  Madsen  showed  that  not 
only  diphtheria  toxin  and  tetanolysin,  but  the  highly  colloidal  diph- 
theria antitoxin  and  antitetanolysin  could  diffuse  into  5  per  cent 
gelatin  jellies. 

Such  a  diffusion  of  colloids  into  a  jelly  naturally  may  be  expected 
if  the  meshes  are  quite  wide,  i.e.,  if  the  jelly  is  quite  dilute. 

Membranes. 

It  will  be  appropriate  to  introduce  the  concept  of  "membrane"  in 
the  following  way:  if  we  make  the  colloid  medium,  the  jelly,  thicker 
and  thicker,  i.e.,  poorer  in  water,  diffusion  must  be  increasingly  hin- 
dered. We  very  soon  reach  a  point  where  no  colloids  are  able  to 
diffuse  into  it  and  we  have  reached  a  special  case  in  our  previous 
exposition,  the  membrane.  We  may  describe  membranes  as  irrever- 
sible gels,  whose  surface  is  very  great  in  relation  to  their  thickness. 
They  play  an  important  role  in  the  organism,  but  we  shall  here 
discuss  their  general  properties  only,  as  their  biological  functions 
will  be  considered  in  Part  III. 

An  excellent  general  resume  with  a  very  complete  bibliography 
has  been  published  by  H.  Zangger  ("Membranes  and  the  Functions 
of  Membrane"). 

On  account  of  the  great  physical  and  chemical  differences  of  the 
membranes  of  the  organisms  the  employment  of  artificial  membranes 
is  preferable  for  the  study  of  their  chief  properties. 

If  a  very  dilute  solution  of  potassium  ferrocyanid  is  carefully 
layered  over  a  concentrated  solution  of  copper  sulphate,  there  is 
formed  at  the  layer  of  contact,  by  chemical  interchange,  a  very  thin 
brown  film  of  copper  ferrocyanid.     Naturally  this  film  is  very  deli- 


PHENOMENA   OF  MOTION  57 

cate  and  is  torn  by  the  slightest  movement.  If  we  add  gelatin  to 
each  solution  and  permit  the  two  salts  to  diffuse  towards  each  other 
in  the  jelly,  there  is  formed  at  the  layer  of  contact  a  very  resistant 
membrane  supported  by  the  jelly.  Expressed  generally,  if  we  per- 
mit two  substances  which  form  a  precipitate  together,  to  diffuse 
towards  each  other  within  a  colloid  medium  which  serves  as  a  sup- 
port, a  membrane  is  formed  at  the  surface  of  contact  which  may 
have,  depending  upon  the  nature  of  the  reacting  substances,  very 
different  degrees  of  permeability. 

Since  the  time  of  Moritz  Traube,*  such  membranes  have  been 
studied,  especially  by  G.  Tammann,*  W.  Pfeffer,*^  Adie,*  P. 
Walden*  and  N.  Pringsheim.*  The  chief  interest,  however,  cen- 
tered in  the  osmotic  phenomena  of  salt  solutions  which  could  be  in- 
vestigated with  the  aid  of  such  precipitation  membranes,  whereas  the 
properties  of  the  membranes  themselves,  with  few  exceptions,  received 
but  secondary  attention.  For  investigations  of  osmosis  the  following 
substances  are  especially  suitable:  ferrocyanid  of  copper  and  ferro- 
cyanid  of  zinc;  indeed  all  ferrocyanid-metal  compounds  are  suitable 
since  they  are  completely  impermeable  to  many  salts.  They  are 
briefly  described  as  semipermeable  membranes,  because  they  are  per- 
meable to  water  though  impermeable  to  most  crystalloids.  If  we 
permit  a  zinc  ferrocyanid  membrane  to  develop  in  a  gelatin  jelly 
and  by  the  addition  of  potassium  ferrocyanid  exercise  a  very  great 
osmotic  pressure,  the  membrane  will  break  in  spite  of  the  jelly  sup- 
port, but  it  will  not  permit  any  potassium  ferrocyanid  solution  to 
diffuse  through. 

Besides  this  extreme  case  there  are  membranes  of  the  most  differ- 
ent permeabilities.  Following  up  the  work  of  the  botanist  N. 
Pringsheim,*  H.  Bechhold  and  F.  Ziegler*^  exhaustively  studied 
such  membranes.  They  impregnated  gelatin  with  silver  nitrate  or 
barium  chlorid,  and  poured  the  molten  solution  into  test  tubes  con- 
taining sodium  chlorid  or  sodium  sulphate.  At  times  a  layer  of 
pure  gelatin  was  interposed.  At  the  surface  of  contact  membranes 
of  silver  chlorid  or  barium  sulphate  were  formed,  which,  however, 
were  permeable  for  the  salt  solution  on  either  side,  because  the  mem- 
branes grew  in  the  direction  of  the  greater  osmotic  pressure,  i.e.,  into  the 
solution  with  the  smaller  osmotic  pressure. 

If,  for  example,  the  silver  nitrate  solution  was  more  concentrated 
it  diffused  through  the  membrane  so  that  the  latter  grew  into  the 
sodium  chlorid  gelatin;  but  if  the  latter  was  more  concentrated  the 
reverse  occurred.  If  both  sides  had  the  same  osmotic  pressure  a  very 
thin  membrane  formed  which  was  sufficient  however  to  arrest  com- 
pletely the  diffusion  of  both  salts.     Evidently  the  meshes  of  the  net- 


58  COLLOIDS  IN  BIOLOGY  AND  MEDICINE  ^ 

work  were  filled  with  membrane-forming  precipitate,  for  as  soon  as 
the  membrane  was  melted,  it  again  became  permeable.  In  the  same 
experiment,  it  was  determined  that  diffusion  was  hindered  only  by 
visible  precipitate  membranes. 

There  is  no  difficulty  in  forming  by  diffusion  similar  precipitate 
membranes  from  pure  organic  materials.  We  have  already,  on  page 
55,  mentioned  that  a  membrane  may  be  formed  by  the  diffusion  of 
goat  serum  into  goat-rabbit  serum  and  we  shall  later  refer  to  the 
fact  that  H.  Bechhold*^  obtained  membranes  by  the  diffusion  of 
metaphosphoric  acid  into  gelatin  containing  albumin.  There  is  no 
doubt  that  they  may  be  obtained  in  other  ways  if  desired.  But  it 
must  by  no  means  be  assumed  that  a  membrane  is  something  rigid 
and  unchangeable;  on  the  contrary,  it  is  constantly  affected  by  the 
substances  which  flow  through  it  and  bathe  it,  making  it  more  or 
less  permeable,  and  in  this  way  under  certain  conditions  may  evoke 
a  self-regulation  or  a  valve-like  action. 

Hitherto  there  have  been  no  investigations  as  to  the  manner  in 
which  the  permeability  of  the  precipitated  membranes  described  is 
influenced  by  crystalloids  diffusing  through  them.  A  priori  it  is  to 
be  assumed  that  such  an  influence  exists  just  as  in  the  case  of  re- 
versible jellies.  That  membranes  may  be  more  or  less  rapidly 
occluded  by  colloids  is  an  observation  which  has  been  frequently 
made  during  the  performance  of  ultrafiltration. 

A  number  of  dried  animal  and  vegetable  membranes  and  parch- 
ment paper  resemble  precipitate  membranes,  in  that  they  possess 
the  same  or  but  slightly  superior  swelling  capacity.  In  dialysis  they 
are  used  for  the  separation  of  colloids  from  crystalloids. 

Most  of  the  membranes  occurring  in  the  organism  are  more  or  less 
swollen;  on  drying  they  lose  this  property  to  a  great  extent,  as  they 
are  inelastic  gels. 

Ultrafilters  (see  p.  95)  formed  by  impregnating  irreversible  jellies 
are  similar  to  natural  membranes,  since  they  must  be  kept  in  water 
to  preserve  their  swollen  condition. 

Membranes  may  be  powerfully  adsorbent,  like  reversible  gels,  and 
in  this  respect  powerfully  influence  diffusion  and  flltration.  Thus 
dyes,  especially  the  basic  ones,  as  well  as  certain  groups  of  enzyms, 
e.g.,  arachnolysin,  staphylolysin  and  rennin  (H.  Bechhold  *4)  are 
strongly  adsorbed  by  many  membranes.  Such  adsorbed  substances 
may  enter  into  chemical  combination  with  the  membrane  (causing 
either  shrinking  or  loss  of  swelling  capacity)  and  thus  diminish  its 
permeability.  This  is  the  effect,  e.g.,  of  tannic  acid,  formaldehyd 
and  Chromates. 

Alcohol,  ether,  acetone  and  sugar  increase  the  permeability  in  cer- 


PHENOMENA   OF  MOTION  59 

tain  low  concentrations;  stronger  solutions  work  to  a  certain  extent 
in  the  opposite  direction. 

The  influence  of  electrolytes  on  the  membranes  of  the  organism  and 
their  permeability  may  depend  on  different  causes.  It  may,  for  ex- 
ample, affect  the  swelling,  and  thus,  the  permeability.  Alkalis  in 
general  increase  the  swelling;  so  also  do  acids  in  great  dilution,  but 
when  concentrated  they  usually  cause  shrinking.  Chemical  changes 
due  to  chromic  acid,  etc.,  diminish  the  permeability. 

The  influence  of  electrolytes  is  not  however  limited  to  this  rather 
indirect  action.  We  shall  see  on  page  77  et  seq.,  that  differences  of 
potential  may  develop  at  the  interface  between  a  solid  phase  and  a 
fluid.  Thus,  for  example,  cellulose  and  wool  are  negatively  charged 
with  respect  to  pure  water.  In  the  case  of  the  majority  of  animal 
membranes,  most  of  which  are  amphoteric,  the  difference  in  potential 
develops  only  in  faintly  alkaline  or  faintly  acid  water.  The  pres- 
ence of  salts  likewise  raises  or  lowers  the  difference  in  potential. 
The  difference  in  the  adsorbability  of  ions  is  accounted  for  also  in 
this  manner.  Wherever  a  difference  in  potential  exists,  the  diffusion 
rate  of  water  is  lowered.  Salts,  on  the  other  hand,  develop  a  differ- 
ence of  potential  in  the  course  of  diffusion;  in  their  passage  through 
a  membrane,  they  raise  or  lower  the  existing  difference  in  potential. 
•Vice  versa,  on  this  account  the  diffusion  rate  of  salts  is  affected  by 
the  difference  in  potential  existing  in  the  membrane. 

A  membrane  may  thus  be  the  seat  of  an  electromotive  force  (F. 
Haber,*  Girard*),  provided  that  it  separates  two  salt  solutions,  or 
a  salt  solution  from  water,  and  that  one  solution  be  faintly  acid  or 
faintly  alkaline.  In  the  first  instance  the  diffusion  through  the  mem- 
brane will  be  much  impeded,  in  the  latter  much  accelerated. 

The  results  of  R.  Burian*^  also  indicate  differences  in  potential  in 
the  ultrafiltration  of  albumin-salt  mixtures;  at  reduced  pressures,  he 
obtained  as  filtrate  a  salt  solution  isotonic  with  the  liquid  filtered. 
If  he  filtered  under  increased  pressure,  the  filtrate  contained  a  lower 
salt  concentration  than  the  original  solution.  F.  G.  Donnan*  has 
made  an  unusually  important  and  fundamental  study  of  membrane 
equilibria,  based  upon  the  osmotic  pressure  and  membrane  potential 
of  electrolytes  containing  a  colloidal  ion.  I  shall  try  to  present 
Donnan's  ideas  without  entering  upon  their  mathematical  basis. 
Let  R  represent  an  acid  colloid,  e.g.,  congo  red,  which  forms  a  salt 
with  Na,  and  let  the  line  which  separates  the  colloid  electrolyte  from 
water  in  our  diagram  be  a  membrane,  impermeable  to  the  colloid. 

(a)  Membrane  hydrolysis.  Let  us  consider  what  occurs  when  the 
outer  water  is  constantly  renewed  as  described  on  page  92  et  seq., 
which  may  be  represented  in  the  following  diagram: 

Initial  condition  Terminal  condition 

RH 


NaOH 


water  RH 


water  NaOH 


60  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

i.e.,  there  is  formed  by  hydrolysis  some  colloid  acid  which  cannot 
diffuse  through,  although  the  NaOH  may  do  so.  If  the  colloid  is  a 
strong  acid,  the  process  terminates  rapidly  providing  the  NaOH  re- 
mains in  the  outer  water,  and  an  osmotic  pressure  may  develop  within 
the  cell  which  is  chiefly  produced  by  the  R  and  Na  components  of  the 
colloid  electrolytes  as  occurs,  for  instance,  in  the  case  of  congo  red. 

If  the  colloid  electrolyte  is  a  weak  acid,  then  proportionally  more 
NaOH  diffuses  outward,  and  when  the  equilibrium  is  established,  it  is 
chiefly  by  the  hydrolytically  split  colloid  acid  and  the  NaOH.  Ex- 
ample: soap  solution. 

If  the  NaOH  is  constantly  removed  (by  continually  renewing  the 
outer  water,  or  by  means  of  a  bond  which  is  not  at  all  or  slightly 
dissociated,  e.g.,  carbonic  acid)  there  will  finally  remain  only  colloid 
acid,  in  fact  the  weaker  the  colloid  acid  the  more  rapidly  the  process 
will  terminate.  What  has  been  stated  for  an  acid  colloid  applies 
of  course  to  a  basic  one  also. 

It  follows  from  these  premises  that  salts  even  of  strong  acids  and 
bases  may  be  completely  broken  up  hydrolytically,  provided  one  ion 
is  a  colloid  which  can  be  held  back  by  a  membrane.  By  membrane 
hydrolysis  it  is  possible  to  separate  from  a  neutral  salt,  either  an 
alkali  (intestinal  or  pancreatic  juice)  or  an  acid  (hydrochloric  acid, 
in  the  stomach,  or  acid  urine).  It  requires  no  special  exposition  to 
show  that  the  same  process  may  be  brought  about  by  ultrafiltration. 

The  reverse  process  may  occur,  however.  If  there  is  a  colloid  acid 
or  base  in  a  cell  surrounded  by  a  membrane,  e.g.,  an  amphoteric 
colloid  albumin  or  fibrin,  a  minimal  concentration  of  H  or  OH  ions 
in  the  outer  fluids  suffice  to  form  a  salt  with  the  colloid  in  the  cell 
which  by  swelling  develops  a  higher  osmotic  pressure. 

(h)  We  shall  now  consider  what  occurs  when  the  colloid  electrolyte 
within  the  membrane  has  an  ion  in  common  with  the  electrolyte 
outside,  e.g.,  the  Na  salt  of  congo  red  (RNa)  and  common  salt  (NaCl). 
We  then  have  the  following  formula: 


Initial  condition 

Equilibrium 

R 

CI 

R 

CI 

Na 

Na 

Na 

Na 

(1) 

(1) 

CI 

(1) 

(2) 

Na  ions  cannot  pass  from  space  (1)  to  space  (2)  since  by  reason  of 
its  colloidal  character  the  anion  R  cannot  follow.^  However,  CI  and 
with  it  the  same  amount  of  Na  will  diffuse  into  (1).     The  amount  of 

1  There  are  always  the  same  number  of  anions  and  cations  in  a  solution.  It 
is  impossible  to  separate  them  by  diffusion  for  then  a  free  electric  charge  would 
be  liberated. 


PHENOMENA   OF  MOTION 


61 


NaCl  which  diffuses  depends  on  the  concentration  of  the  solutions 
in  space  (1)  and  space  (2).  If  the  concentration  in  space  2  (C2)  is 
high  in  proportion  to  that  in  space  1  (Ci)  much  NaCl  will  pass  from 
(1)  to  (2)  and,  if  the  conditions  are  reversed,  only  a  little  will  do  so. 
Mathematically  represented  (assuming  complete  electrolytic  dis- 
sociation) we  have  the  following  equation,  where  Ci  or  C2  represent 
the  molar  ion  concentration  in  the  respective  spaces  and  X  the  frac- 
tion of  the  molar  ion  concentration  which  diffuses  from  (2)  to  (1) : 

C2  —  -A       Ci  -7-  Co 


X 


Co 


X     c 

If  C2  is  small  in  comparison  with  Ci  we  may  express  it :  yr  =  tt' 

C2      Ci 

X      1 

If  Ci  is  very  small  the  equation  becomes  jr  =  ?;' 

C2      2i 

The  following  table  taken  from  Donnan's  work  illustrates  the  dis- 
tribution of  NaCl. 


Original  concentration 
of  NaR  in  (1), 

Original  concentration 
of  NaCl  in  (2), 

Original  relation  of  NaR 
to  NaCl, 

C2 

Per   cent    NaCl    dif- 
fusing from  (2)  to  (1), 
100  X 

0.01 
0.1 

1 
1 
1 

1 
1 
1 

0.1 
0.01 

0.01 
0.1 

1 

10 

100 

49.7 
47.6 
33 

8.3 

1 

Though  we  might  assume  d  priori  that  the  NaCl  would  distribute 
itself  equally  in  both  spaces  in  the  presence  of  a  membrane  absolutely 
permeable  for  it,  this  table  shows  that  the  colloid  electrolyte  has  a 
remarkable  influence  as  soon  as  the  concentration  of  NaCl  falls.  To 
a  certain  extent  the  colloid  electrolytes  drive  the  NaCl  out  of  the 
cell.  If  Ci  =  1  only  about  11  per  cent  of  a  physiological  salt  solu- 
tion (C2  =  0.145)  could  penetrate  the  cell;  or  if  it  were  already  in 
the  cell  it  was  reduced  to  about  11  per  cent.  Apparently  the  mem- 
brane is  permeable  only  from  one  side  for  the  readily  diffusible  NaCl. 
(c)  Finally  we  must  consider  the  case  when  the  colloid  electrolyte 
in  the  membrane  is  opposed  to  an  electrolyte  without  an  ion  in  com- 
mon, as  for  instance: 


Initial  condition 

Equilibrium 

Na 

K 

Na 

K 

R 

CI 

K 

Na 

(1) 

(2) 

CI 
R 

CI 

(1) 

(2) 

62  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Na  will  diffuse  out,  K  and  CI  will  diffuse  in.  Thus  we  obtain  the 
following  equation  if  CNa(i)  expresses  the  molar  concentration  of 
Na  in  space  (1). 

CNa(l)  _  Ck(1)   _  Cci(l)   _  Ci  +  C2  _ 
CNa(2)  Ck(2)         Cc1(2)  Ca 

If  the  concentration  in  the  cell  (Ci)  is  large  compared  with  the  outer 
solution  (C2),  then 

Ci  -|-  C2  _  Ci 
C2  C2 

If  Ci  is  small,  then 

Ci  +  C2  _  . 
C2 

Let  us  consider  the  equation  representing  a  condition  that  frequently 
occurs  physiologically.  If  Ci  =  100  and  C2  =  1  it  follows  that  99 
per  cent  of  the  Na  originally  present  in  (1)  will  remain  in  (1) ;  only 
1  per  cent  will  diffuse  into  (2).  99  p3r  cent  of  the  K  originally  pres- 
ent in  (2)  will  diffuse  into  (1);  only  1  per  cent  of  the  CI  originally 
present  in  (2)  will  diffuse  into  (1). 

From  this  we  may  understand  the  hitherto  inexplicable  distribution 
of  salt  in  cells,  e.g.,  in  red  blood  corpuscles.  A  colloidal  anion  attracts 
the  foreign  crystalloid  cation  and  drives  out  the  anions.  Colloidal 
cations  act  in  the  reverse  way. 

In  conclusion  Donnan  derived  formulas  for  the  differences  in 
electric  potential  which  must  exist  after  the  equilibria  described 
have  been  reached  (membrane  potentials). 

There  are  already  many  theories  to  explain  the  difference  in  poten- 
tial in  organs  and  the  electrical  currents  which  occur  in  the  body 
(muscle,  nerve,  electric-fish).  These  theories  have  the  error  that 
they  require  conditions  which  do  not  exist  in  the  body,  and  in  part 
that  much  greater  differences  in  potential  arise  in  the  body  than 
would  be  possible  according  to  these  theories.  In  this  respect 
Donnan's  theory  differs  much  from  its  predecessors,   f 

We  shall  not  present  his  formula  here  but  only  a  general  explana- 
tion: If  there  are  two  equally  concentrated  solutions  of  NaCl  sep- 
arated by *ä -membrane,  and  we  insert  a  piece  of  platinum  foil  into 
each  and  connect  them  with  a  wire,  no  current  will  flow  in  the  wire. 
If  the  solutions  are  of  different  concentrations,  theoretically,  electric 
energy  wih  be  evident  until  the  differences  in  concentration  disappear 
as  the  result  of  diffusion.  Such  systems  are  called  "concentration 
couples." 

A  sy'stern  consisting  of  colloid  electrolyte,  salt  or  water  is  a  con- 
centration couple  which  develops  a  current  in  passing  from  "initial 
condition"  to  "equilibrium."     This  may  be  ascribed  to  the  unequal 


PHENOMENA   OF  MOTION  63 

concentration  of  ions  on  the  two  sides  of  the  membrane.  Let  us 
consider  the  simplest  illustration  of  the  equilibrium  between  NaR 
and  NaCl  represented  in  our  table  on  page  60  where  the  original  con- 
centration is  NaR  :  NaCl  =  1:1  and  equihbrium  is  established  when 
33  per  cent  NaCl  passes  from  (2)  to  (1). 
The  schematic  representation  would  be: 


Equilibrium 

Na 

Na 

Na 

Na 

CI 

CI 

R 

CI 

(1) 

(2) 

In  this  instance  all  charges  are  mutually  satisfied  excepting  those  of 
CI  and  R.  The  slowly  diffusing  anion  R  is  opposed  to  the  rapidly 
diffusing  anion  CI  so  that  a  difference  of  potential  must  arise  at  the 
boundary  surface.  In  the  cases  hitherto  described,  the  membrane 
itself  is  the  seat  of  the  difference  in  potential.  The  conditions  are 
quite  different  if  the  membrane  acts  only  as  a  bounding  surface,  that 
is,  if  it  is  not  equally  permeable  for  all  ions.  In  this  case,  the  uni- 
versally present  "contact  potential,"  existing  in  two  contiguous 
salt  solutions,  is  modified  by  the  aforementioned  property  of  the 
membrane. 

Finally,  we  must  recall  another  kind  of  membrane  which  does  not 
fall  into  any  of  the  previous  categories.  According  to  W.  Nernst, 
a  film  of  water  upon  ether  forms  a  semipermeable  membrane  for 
benzol.  The  experiment  is  carried  out  in  this  way:  A  pig's  bladder 
is  soaked  in  water;  the  bladder  plays  no  part,  other  than  to  hold  the 
water  which  forms  a  partition  between  ether  and  ether  containing 
benzol.  Here  the  semipermeability  of  the  membrane  depends  en- 
tirely on  selective  solubility.  Benzol  is  insoluble  in  water;  ether  on 
the  contrary  has  a  limited  solubility,  and  as  a  result  ether  diffuses 
through  the  water  to  the  benzol.  Subsequently  many  such  com- 
binations were  devised.  They  are  very  extensively  distributed  in  the 
organism.  It  is  unnecessary  to  think  of  complete  semipermeability 
in  every  case;  scattered  deposits  (fat,  lecithin,  etc.)  nraj^^suffice  to 
bring  about  a  partial  permeability. 

The  membranes  of  Wistinghausen  depend  on  this  principle  of 
selective  permeability.  He  impregnated  with  gallic  acid  salts, 
animal  membranes  which  then  became  permeable  for  fat;  by  merely 
washing  away  the  salts  the  permeability  is  abolished.  Attention 
should  be  called  to  a  remarkable  observation  of  Zott  (cited  by 
H,  Zangger)  which  belongs  in  this  chapter.  He  discovered  that  a 
membrane  through  which  sugar  has  diffused,  permitted  the  passage 
of  gum  arable  after  it  had  been  moistened  with  alcohol. 


CHAPTER  V. 

CONSISTENCY  OF  COLLOIDS. 

Internal  Friction. 

The  various  colloids  show  all  possible  transitions  from  fluids  to 
solid  substances.  A  fluid  may  take  on  any  shape,  and  the  work 
necessary  to  change  its  form,  i.e.,  to  overcome  its  internal  friction,  is 
very  slight.  Solid  substances,  according  to  Wilhelm  Ostwald,  pos- 
sess a  form-energy  also  called  elasticity;  the  energy  necessary  to 
change  their  form,  i.e.,  to  overcome  their  internal  friction,  is  very 
great.  If  we  picture  to  ourselves  a  number  of  colloids  and  gels  we 
pass  from  a  true  fluid,  water  for  instance,  through  the  albumoses 
and  albumin  solutions  to  the  semifluid  gels  {e.g.,  1  per  cent  gelatin), 
jellies  and  finally  to  the  firm  substances  {e.g.,  horn). 

High  internal  friction,  viscosity,  is  a  typical  property  of  hydrophile 
colloids.  Since  colloids  are  diphasic  systems,  the  internal  friction 
will  depend,  above  all,  upon  the  size  of  the  free  surface  of  the  colloid, 
i.e.,  upon  the  concentration.  Changes  in  temperature  are  of  great 
importance.  The  absolute  as  well  as  the  relative  influence  of  concen- 
tration is,  indeed,  characteristic  for  colloids.  Even  traces  of  colloids 
(agar,  gelatin)  may  increase  the  viscosity  of  water  to  an  extraordi- 
nary extent.  We  may  obtain  all  degrees  of  internal  friction  with 
agar  and  in  fact  a  5  per  cent  solution  of  agar  is  a  solid  body  at  room 
temperature. 

Usually  the  viscosity  increases  with  decrease  in  temperature,  inas- 
much as  substances  then  approach  the  solid  condition.  Gelatiniza- 
tion  is  analogous  to  the  sohdification  of  a  molten  fluid,  where  internal 
friction  rapidly  rises  within  a  small  temperature  range. 

J.  Friedländer,*  D.  Holde*  and  V.  Rothmund*  proved  that 
artificial  emulsions  (gum  water,  castor  oil,  so-called  solid  fats)  exhibit 
a  variation  in  their  viscosity  curves  according  to  temperature  and 
concentration,  similar  to  that  shown  by  many  natural  hydrophile 
colloids.  T.  B.  Robertson*  found  that  emulsions  of  oil  in  water 
became  increasingly  more  viscous  the  higher  the  concentration  of 
the  oil,  until  a  critical  point  was  reached  when  the  viscosity  decreased; 
the  water  then  became  the  dispersed  phase. 

Internal  friction  is  indeed  a  very  complicated  phenomenon.  It 
depends  according  to  W.  B.  Hardy  upon  (1)  the  internal  friction  of 

64 


CONSISTENCY  OF  COLLOIDS  65 

the  different  phases,  (2)  the  surface  friction  of  the  internal  surfaces, 
(3)  the  surface  tension  of  the  internal  surfaces  and  (4)  the  strength 
of  the  electrical  charge.  To  what  extent  the  individual  factors  influ- 
ence the  internal  friction  is  as  yet  unknown. 

We  have,  however,  received  valuable  guidance  from  the  study  of 
the  effect  of  electrolytes.  Especially  remarkable  is  the  parallelism 
between  swelling  and  internal  friction.  It  depends,  apparently,  on 
the  fact  that  both  phenomena  are  characterized  by  an  increase,  i.e., 
multiplication,  of  the  free  surfaces.  Thus,  for  instance,  we  see  that 
acids  and  alkalies  which  favor  the  swelling  of  gelatin  also  increase 
the  internal  friction  of  albumin,  because  there  probably  occurs  an  in- 
crease in  the  free  surfaces  of  the  albumin  ions  (see  p.  153  et  seq.). 

Swelling  and  Shrinking. 

If  a  crystalloid  (common  salt,  sugar)  is  thrown  into  water,  it  sub- 
divides in  it  until  finally  it  is  completely  dissolved;  the  particles  of 
salt  or  sugar  lose  their  cohesion.  A  colloid  (glue,  wood)  in  contact 
with  water  increases  its  volume,  it  swells;  its  particles  retain  their 
cohesion.  This  property,  however,  is  possessed  only  by  hydrophile 
colloids. 

The  imbibition  of  water,  that  is,  swelling,  may  either  go  on  in- 
definitely in  the  case  of  colloids,  so  that  finally  the  particles  are  torn 
asunder  and  a  solution  or  sol  is  formed  as  in  the  case  of  albumin;  or 
the  imbibition  may  reach  its  limit  very  rapidly,  as  in  the  case  of  wood. 
Between  these  there  are  all  sorts  of  transitions,  e.g.,  glue.  Only  in 
the  case  of  gels  is  it  usual  to  refer  to  swelling.  In  the  organism,  gels 
having  very  slight  ability  to  swell  serve  as  covering  and  framework 
{e.g.,  hide,  collagen,  shells  and  wood).  They  are  intended  to  retain 
the  outward  form.  The  same  is  true  of  the  supporting  tissues  of 
the  individual  organs  and  even  of  the  cells,  vessel  walls,  the  mem- 
branes of  the  intestinal  canal,  connective  tissues,  the  vascular  bundles 
of  plants,  cell  membranes,  etc.  On  the  other  hand,  the  cell  content 
possesses  the  ability  to  swell  to  a  high  degree. 

Every  organ  has  a  certain  definite  normal  fluid  content.  A  healthy 
plant  has  a  definite  turgescence  and  the  protoplasm  of  a  healthy 
animal  a  given  degree  of  swelling;  every  abnormal  change  in  this 
signifies  illness  or  even  death.  Without  doubt  swelling  plays  a  very 
important  role  in  the  case  of  many  phenomena  which  have  hitherto 
been  attributed  to  osmotic  pressure.  Indeed,  the  osmotic  pressure 
is  only  manifested  completely  in  the  presence  of  a  semipermeable 
membrane,  whereas  the  ability  to  swell  does  not  require  the  presence 
of  a  membrane.  Swelling  may  under  some  circumstances  counter- 
balance the  osmotic  pressure  or  even  overcome  it  and  concentrate 


66 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


solutions.  An  excellent  example  of  the  latter  is  described  by  C. 
Ludwig.  He  hung  a  well-dried  animal  bladder  in  concentrated  salt 
solution.  The  bladder  swelled,  taking  up  a  dilute  salt  solution  and 
common  salt  crystallized  out  in  the  remainder.  Amphibia,  e.g., 
frogs,  may  lose  one-fourth  of  their  body  weight  upon  drying  as  has 
been  shown  by  E.  Overton.*^  Although  they  contain  about  80 
per  cent  water,  the  osmotic  pressure  of  the  blood  almost  doubles. 
The  explanation  of  this  is  that  only  a  portion  is  water  of  solution, 
the  remainder  is  water  of  swelling.  Upon  diying  the  water  of  swell- 
ing is  more  strongly  retained  than  the  water  of  solution. 

Swelling  exhibits  manifestations  of  energy  to  no  less  a  degree  than 
osmotic  pressure.  I  shall  give  several  examples  taken  from  Wolf- 
gang Ostwald's  "Grundriss."  According  to  the  investigations  of 
the  plant  physiologist,  Hales,  swelling  peas  were  able  to  lift  the 
cover  of  an  iron  pot  weighted  with  83.5  kilograms.  H.  Rodewald 
found  that  it  requires  2523  atmospheres  pressure  to  overcome  the 
swelling  pressure  of  starch.  J.  Reinke*  determined  the  swelling 
pressure  of  laminaria,  a  sea  weed.  Some  of  his  data  quoted  from 
H.  Freundlich  give  us  an  idea  of  the  enormous  pressures,  changes 
in  volume  and  amounts  of  water  taken  up  when  swelling  occurs 
and  of  the  pressures  required  for  dehydration.  Ten  layers  of  dried 
laminaria  scales  each  0.1  mm.  thick  and  50  mm.^  were  placed  in  the 
apparatus. 


Pressures  in  atmos- 
pheres. 

Elevation  in  mm.  due 
to  swelling. 

Pe-centage  of  water 

by  volume  in 
air-dried  substance. 

41.2 
21.2 

7.2 

1 

0.16 
0.35 
0.97 
3.30 

16 

35 

97 

330 

We  obtain  a  fair  idea  as  to  the  general  course  of  swelling  by  ob- 
serving a  sheet  of  gelatin.  Dry  gelatin  takes  up  one-third  of  its  weight 
of  water  from  a  moisture-saturated  atmosphere  at  room  temperature, 
in  order  to  reach  a  condition  of  equilibrium.  If  this  sheet  is  then 
placed  in  a  dish  of  cold  running  water  it  absorbs  from  it  10  times  its 
dry  weight  of  water  in  order  again  to  reach  a  condition  of  equilibrium. 
In  dry  air  the  water  evaporates  and  shrinking  occurs.  The  experi- 
ment may  be  repeated  as  often  as  desired  with  the  same  result.  On 
this  account  substances  of  this  group  are  termed  elastic  gels. 

Coagulated  albumin,  e.g.,  boiled  fibrin,  behaves  differently.  If  it 
is  air-dried  a  horny  residue  remains  which,  though  it  takes  up 
some   water,   or  swells  when  it  is  placed  in  water,  never   again 


CONSISTENCY  OF  COLLOIDS  67 

approaches  its  original  gelatinous  state;  such  gels  are  called  inelastic 
gels.  One  of  the  most  important  operations  of  microscopical  technic 
is  the  hardening  of  elastic  gels.  (See  Chapter  XXIII.)  Their  ability 
to  swell  in  water  is  destroyed  by  the  chemical  action  of  formalin, 
chromic  acid,  mercuric  chlorid,  etc.  We  must  consider  that  in  the 
organism,  the  inelastic  gels,  i.e.,  connective  tissue  and  also  the  cell 
pellicle,  etc.,  arise  from  elastic  gels  by  chemical  changes  with  con- 
sequent loss  of  water  or  drying,  as  may  be  observed  at  the  surface  of 
any  wound.  In  this  cormection,  we  may  revert  to  the  formation 
of  surface  films  (see  p.  33),  whose  formation  is  certainly  more  than 
merely  analogous  to  that  of  organized  membranes,  skin,  etc. 

Classical  studies  on  the  swelling  and  shrinking  of  slightly  elastic 
gels  were  made  by  J.  M.  van  Bemmelen  in  the  case  of  silicic  acid 
gel  and  amplified  by  0.  Bütschli,  So  many  difficulties  are  unfor- 
tunately offered  to  the  application  by  analogy  of  these  properties 
to  organized  inelastic  gels,  that  we  must  confine  our  attention  to 
the  most  important  ones.  The  evaporation  of  water  from  a  silicic 
acid  gel  proceeds  at  first  as  it  would  from  a  solution.  When  the  gel 
reaches  a  certain  consistency  a  turbidity  appears,  that  is,  hollow 
spaces  of  about  o  fifj,  form  between  the  supporting  walls  of  the  gel 
which  become  filled  with  air.  Upon  losing  still  more  water,  the  tur- 
bidity disappears  and  the  gel  becomes  glassy.  In  this  latter  respect 
the  inelastic  gel  of  silicic  acid  differs  very  materially  from  the  elastic 
gel  of  gelatin,  which  does  not  become  turbid.  This  is  likewise  the 
case  in  the  reabsorption  of  water.  Though  gelatin  shows  a  similar 
curve  both  on  swelling  and  shrinking,  silicic  acid  gel  and  indeed  we 
may  say  all  inelastic  gels  show  entirely  different  curves.  That  is,  the 
swelhng  of  elastic  gels  is  practically  completely  reversible,  whereas 
with  inelastic  gels  this  is  not  the  case. 

The  changes  a  gel  undergoes  on  freezing  and  thawing  are  very 
similar  to  those  of  shrinking  and  swelling.  The  crystallization  of  ice 
from  a  gel  containing  water  indicates  a  withdrawal  of  water,  whereas 
upon  thawing,  water  becomes  again  available  for  swelling  (H.  W. 
Fischer,  0.  Bobertag  and  C.  Feist*).  There  are  consequently 
substances  which  after  freezing  and  thawing  revert  almost  completely 
to  their  original  state,  e.g.,  soluble  starches,  fish  glue,  whereas  others, 
e.g.,  silicic  acid  hydrosol  and  albumin,  undergo  changes  which  are 
more  or  less  irreversible. 

The  influence  of  electrolytes  on  the  swelling  of  gelatin,  agar,  pig's 
bladder,  cartilage  and  fibrin,  is  very  considerable.  It  has  been  in- 
vestigated especially  by  F.  Hofmeister,*  Wo.  Pauli, *i  K.  Spiro,* 
Wo.  OsTWALD,*^  and  Martin  H.  Fischer.*  It  may,  in  general,  be 
stated  that  acids  and  alkalis  increase  the  swelling  capacity  to  an 


68 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


extraordinary  degree.  This,  however,  does  not  depend  only  upon 
the  electrolytic  dissociation  of  various  acids  and  the  concentration  of 
H  or  OH  ions.  In  the  case  of  strong  acids  it  reaches  a  maximum 
at  a  certain  concentration  and  then  decreases.  Thus  Martin  H. 
Fischer  found  that  fibrin,  which  swelled  to  8  mm.  in  water,  in 
0.02  normal  HCl  reached  the  maximum  swelling  of  48  mm. ;  whereas 
in  0.1  normal  HCl  the  swelling  reached  only  21  mm.  In  the  case  of 
H2SO4  the  maximum  swelling  was  only  11  mm.  in  0.024  normal  acid. 
Purified  glutin  (according  to  the  experiments  of  R.  Chiari  in  Pauli 's 


Fig.  7.    The  swelling  of  fibrin  in  solutions  of  various  sodium  salts 
(^V  molecular).     (From  M.  H.  Fischer.) 

laboratory)  is  so  sensitive  to  acids  that  it  swells  less  in  distilled 
water  than  in  Vienna  Hochquellwasser,  because  of  the  CO2  contained 
in  the  latter.  Furthermore,  distilled  water  may  even  be  distin- 
guished from  conductivity  water  by  swelling  experiments  with 
glutin.  The  swelling  in  alkalis  is  still  greater;  in  0.02  normal  NaOH 
it  reached  77  mm.  M.  H.  Fischer  believes  that  the  swelling  in 
acids  is  dependent  upon  the  concentration  of  the  H  ions  minus  the 
effect  of  the  anions  of  the  acid  under  consideration.  In  this  case, 
there  probably  exists  an  antagonism  between  cation  and  anion,  such 
as  may  be  demonstrated  in  the  case  of  neutral  salts.  A  similar  rule 
probably  obtains  for  alkalis. 


CONSISTENCY  OF  COLLOIDS  69 

When  such  heterogeneous  substances  as  gelatin,  fibrin,  etc.,  behave 
similarly  under  the  influence  of  electrolytes,  we  may  assume  that  the 
same  cause  determines  the  behavior,  and  this  cause  must  be  sought 
in  the  chemical  nature  of  these  substances.  Apparently  these  sub- 
stances are  amphoteric,  i.e.,  at  the  same  time  weak  acids  and  weak 
bases,  forming  under  the  influence  of  acids  and  bases,  more  or  less 
ionized  salts.  Ionization  causes  an  hydration,  i.e.,  an  imbibition  of 
water  which  is  evidenced  in  the  case  of  these  gels  by  their  capacity 
to  swell,  and  in  the  case  of  dissolved  albumins,  by  an  increase  of  the 
internal  friction.  We  should  therefore  not  expect  to  see  these  phe- 
nomena in  the  case  of  gels  of  entirely  different  chemical  properties, 
silicic  acid  gel,  for  example. 

Neutral  salts,  to  a  certain  extent,  favor  the  imbibition  of  water  and 
indeed  the  swelling  is  greater  in  such  dilute  salt  solutions  than  it  is 
in  pure  water.  At  a  certain  concentration  (for  NaCl,  13.8  per  cent) 
the  amount  of  fluid  taken  up  reaches  a  maximum  and  then  falls 
again.  The  anions  are  primarily  active  in  favoring  swelling,  whereas 
the  cations  have  a  lesser  influence  and,  in  favoring  swelling, 

CNS  >  I  >  Br  >  NO3  >  CIO3  >  CI; 

whereas  in  favoring  shrinking, 

SO4  >  tartrate  >  citrate  >  acetate. 

Although  a  dry  jelly  removes  more  water  from  a  salt  solution  than 
it  does  salt  so  that  the  concentration  of  the  solution  is  increased,  a 
swollen  jelly  takes  up  more  salt  than  it  does  water,  thus  diminishing 
the  concentration  of  the  solution.  The  swelling  in  acid  or  alkaline 
solution  is  always  much  decreased  hy  the  presence  of  neutral  salts, 
and  anions  are  much  more  active  than  cations.  In  producing  this 
decrease 

citrate  >  tartrate  >  phosphate  (?)  >  SO4  >  acetate  >  I  >  CNS 

>N03>Br>Cl; 

Fe  >  Cu  >  Sr  >  Ba  >  Ca  >  Mg  >  NH4  >  Na  >  K. 

Though,  e.g.,  0.78  gm.  gelatin  in  100  cc.  of  0.05  nHCl  swelled  up  to 
14.61  gm.,  the  swelling  reached  only  2.84  gm.  in  the  presence  of  -^ 

potassmm  citrate  and  about  7  gm.  in  the  presence  of  -^  KCl. 

Antagonistic  Effects.  In  addition  to  the  antagonistic  action  of  neu- 
tral salts  on  the  swelling  due  to  H  and  OH  ions,  there  is  also  an 
antagonism  to  monovalent  cations  hy  'polyvalent  cations. 


70  COLLOIDS  IN  BIOLOGY  AND  MEDICINE  _ 

Thus  it  has  been  determined  by  the  experiments  of  Martin  H. 
Fischer  on  fibrin  and  of  Wo.  Ostwald  on  gelatin  that  swelHng  is 
much  more  strongly  depressed  by  polyvalent  cations  than  by  mono- 
valent ones  (Mg  <  Ca  <  Ba  <  Sr  <  Cu  <  Fe)  and  it  seems  probable 
that  polyvalent  cations  counteract  the  action  of  monovalent  ones  in 
favoring  swelling.  So  far  as  I  know  there  is  as  yet  no  colloid  chemi- 
cal confirmation  of  this  assumption  in  the  case  of  swollen  colloids. 
There  are,  however,  a  number  of  biological  experiments  concerning 
the  inhibition  of  the  poisonous  action  of  neutral  salts  by  polyvalent 
cations  (see  p.  378),  which  in  all  probability  are  referable  to  the  in- 
hibition of  harmful  swelling.  According  to  these  biological  experi- 
ments the  antitoxic  effect  of  cations  increases  with  their  valence  and 
stands  in  relation  to  the  ionization  pressure  or  the  electrolytic  solu- 
tion tension.^ 

Our  present  knowledge  indicates  that  the  swelling  and  the  shrink- 
ing of  hydrophile  gels  absolutely  parallels  the  formation  of  ions  and 
neutral  particles  in  the  case  of  albumin.  This  has  been  more  ex- 
haustively discussed  on  page  153  et  seq.  The  same  factors  which  favor 
the  ionization  of  albumin,  namely,  acids  and  alkalies,  also  favor 
swelling.  In  this  case  as  in  the  other  the  presence  of  neutral  salts 
depresses  the  action  of  acids  and  alkalies,  polyvalent  cations  or 
anions  acting  more  powerfully  than  monovalent  ones.  We  rec- 
ognize in  both  ionization  and  swelling  a  tendency  towards  an  in- 
crease of  the  free  surface,  which  is  associated  with  the  taking  up  of 
water.  This  may  go  so  far  that  the  molecules  are  split;  hydrolysis 
occurs  and  cleavage  products  are  formed.  Accordingly,  chemical  re- 
actions, especially  hydrolytic  cleavages,  occur  much  more  rapidly 
in  swollen  than  in  shrunken  colloids.  For  instance  (according  to 
E.  Knoevenagel*),  the  hydrolysis  of  swollen  acetyl-cellulose  by 
potassium  hydrate  requires  only  a  few  minutes;  but  the  same  process 
requires  days  in  the  case  of  the  shrunken  material. 

Non-electrolytes  have  only  a  slight  influence  on  swelling.  Of  the  few 
cases  known  to  us  we  may  mention  that  urea  favors  the  swelling  of 
gelatin  even  in  acid  solution,  but  it  has  no  effect  on  fibrin.  Alcohol 
and  sugar  favor  the  swelling  of  gelatin  in  a  certain  concentration  be- 
tween 1  and  2  per  cent. 

These  data  were  almost  exclusively  obtained  with  gelatin  and 
fibrin.  Both  gels  behave  qualitatively  alike  (excepting  with  urea) 
though  there  are  quantitative  differences.     Fibrin  swells  much  more 

^  To  avoid  any  misunderstanding  it  should  be  stated  that  substances  which 
are  themselves  strongly  toxic,  e.g.,  barium,  zinc  or  lead  salts,  may  act  in  suit- 
able small  doses  as  antidotes  (probably  by  counteracting  swelling)  to  harmful 
quantities  of  neutral  solutions  {e.g.,  pure  NaCl  solutions). 


CONSISTENCY  OF   COLLOIDS  71 

than  gelatin.  Gelatin  may  absorb  about  25  times  its  weight  in 
water;  fibrin  40  times.  The  order  in  which  neutral  salts  act  on 
gelatin  is  different  from  that  in  which  they  act  upon  fibrin. 

It  may  be  assumed  that  the  different  gels  of  the  organism  vary 
quantitatively  in  their  behavior  under  the  influence  of  the  same  elec- 
trolytes and  it  is  obvious  that  the  salt  absorption  of  different  gels 
varies  as  well  as  their  water  absorption.  An  investigation  of  the 
water  and  salt  absorption  of  different  kinds  of  gels  in  the  presence  of 
mixtures  of  electrolytes  is  much  to  be  desired.  Our  knowledge  of 
tissues  and  secretions  forces  us  to  the  conclusion  that  the  different 
tissues  possess  a  very  different  specific  ability  to  absorb  certain  sub- 
stances or  ions.  Only  thus  can  we  understand  why  the  blood  cor- 
puscles withdraw  more  potassium  salts  from  the  lymph,  the  cartilages 
more  sodium  salts  and  the  bone-building  tissues  more  calcium  salts. 
Only  thus  can  we  obtain  a  conception  of  the  specific  crystalloid  con- 
tent of  various  secretions  and  of  selective  resorption. 

The  Crystallization  of  Colloids. 

Though  P.  P.  VON  Weimarn  describes  the  crystalline  state  as  the 
"sole  ultimate  condition  of  matter"  which  is  characteristic^  for  all 
substances  (even  gases),  we  shall  not  attempt  here  a  critical  study  of 
this  theory  nor  determine  the  limits  of  the  crystallization  of  solids. 
We  shall  consider  only  how  crystals  occur  in  colloidal  substances 
and  more  particularly  limit  ourselves  to  the  biocolloids.  We  know 
only  a  limited  number  of  crystallizable  biocolloids;  the  most  im- 
portant are  egg  albumin,  horse  serum  albumin,  certain  plant  albumins 
(aleuron  crystals  from  Para  nuts,  cotton,  hemp  and  sunflower  seeds), 
oxyhemoglobins,  hemoglobin  and  methemoglobin.  Egg  albumin  has 
been  obtained  in  the  shape  of  needles,  the  albumins  of  vegetable  seeds 
partly  in  octahedra  and  partly  in  tablet-shaped  hexagonal  prisms. 
Oxyhemoglobins  crystallize  in  various  ways  depending  upon  the 
animal  species  from  which  they  are  derived.  For  example,  horse 
oxyhemoglobin  forms  rhombic,  and  squirrel  oxyhemoglobin  forms 
hexagonal  prisms.  These  substances  may  be  recrystallized  and  under 
the  same  conditions  give  the  identical  crystal  form.  It  may  be  re- 
marked in  passing,  that  many  crystals  giving  the  albumin  reaction 
have  frequently  been  observed  in  organs,  but  they  have  been  in- 
sufficiently studied.  [Crystalline  form  is  markedly  influenced  by  the 
presence  of  protective  colloids  in  the  crystallizing  solution.  See  J. 
Alexander,  Kolloid  Zeitschrift,  iv,  p.  86.  Tr.]  Crystalline  products 
have  been  obtained  from  starches,  e.g.,  sphero-crystals  from  inulin. 

^  Bibliography  given  in  Wo.  Ostwald's  Grundiss  der  Kolloidchemie  (Dresden, 
1911). 


72  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  crystallization  of  alkaline  salts  of  the  higher  fatty  and  arylic 
acids  is  well  known. 

The  crystals  of  colloids  are  distinguishable  from  those  of  crystal- 
loids in  many  respects.  That  their  solution  is  preceded  by  a  swelling 
is  not  surprising  in  view  of  the  hydrophile  colloidal  character  of  the 
substances  under  consideration.  On  the  other  hand,  it  is  remarkable 
that  other  constituents  may  always  be  demonstrated  as  inclusions. 
The  crystallized  globulins  from  vegetable  seeds  always  contain  com- 
mon salt.  K.  A.  H.  MoRNER*  showed  that  only  the  sulphates  of 
egg  and  serum  albumin  were  crystallizable.  As  E.  Abderhalden*^ 
has  shown,  oxyhemoglobin  crystals  do  not  contain  their  proper  pro- 
portion of  albumin,  and  although  numerous  researches  on  crystallized 
egg  albumin  have  been  undertaken,  in  different  instances  the  amount 
of  contained  carbohydrate  varied. 

In  spite  of  these  facts,  we  are  of  the  opinion  that  colloids  can 
actually  crystallize,  and  that  their  crystal  form  is  not  controlled  by 
the  crystalloid  impurities.  We  know  that  crystalloids  frequently 
include  mother  liquor,  that  they  may  form  mixed  crystals  and 
that  it  is  often  impossible  to  remove  impurities  by  ordinary  re- 
crystallization.  In  view  of  the  persistent  salt  content  of  crystallized 
albumins  it  is  probable  that  only  their  salt-like  compounds  possess  a 
definite  crystalline  shape.  Especially  favorable  to  this  view  is  the  fact 
reported  by  Dabrowski,  that  crystallized  egg  albumin,  when  placed 
in  a  3.6  per  cent  solution  of  ammonium  sulphate,  exhibits  a  more 
rapid  diffusion  than  salt  free  egg  albumin,  and  has  about  one-sixth 
of  the  atomic  volume  of  the  latter.  The  crystallized  egg  albumin, 
therefore,  is  formed  of  smaller  particles. 

The  Life  Curve  of  Colloids. 

Though  in  the  absence  of  chemical  changes,  crystalloids  retain  their 
physical  properties,  in  the  case  of  colloids  after  a  lapse  of  time  changes 
occur  which  are  conunonly  called  aging.  For  instance,  silicic  acid 
which  has  been  freshly  prepared  from  water-glass  solution  and  HCl 
is  at  first  dialyzable  but  loses  this  property  after  a  few  days.  Most 
of  the  "aging  phenomena"  of  sols  are  characterized  by  the  fact  that 
the  particles  of  a  highly  dispersed  solution  gather  together  to  form 
larger  particles,  that  their  sensitiveness  to  flocculation  is  increased 
or  that  they  spontaneously  coagulate.  In  the  case  of  gels,  their 
elasticity  suffers  changes  and  they  become  optically  inhomogeneous 
or  turbid. 

Bearing  in  mind  that  colloids  are  metastable  systems,  it  is  obvious 
that  in  the  course  of  time  they  must  change,  since  they  tend  to  be- 
come stable  systems.    In  the  examination  of  a  colloid,  the  properties 


CONSISJ'ENCY  OF  COLLOIDS  73 

found,  strictly  speaking,  are  applicable  to  its  momentary  condition; 
previously  and  subsequently  it  has  different  properties.  Every  point 
of  its  life  curve  has  a  previous  history  and  the  final  portions  of  this 
constantly  flattening  curve  are  the  aging  phenomena.  In  con- 
tradistinction to  crystalloids  every  colloid  is  a  particular  individual. 
If  solutions  of  hydrophobe  colloids,  e.g.,  arsenic  sulphid,  gold  solu- 
tion, etc.  (without  protective  colloid),  are  permitted  to  stand  for 
some  time,  they  flocculate  after  a  short  time  or  else  after  a  lapse 
.  of  years.  It  may  be  that  traces  of  electrolytes  are  responsible 
for  the  flocculation.  In  other  cases,  electrolytes  certainly  play  no 
part;  as  I  shall  show  by  a  number  of  examples,  there  is  an  evident 
tendency  for  the  unstable  colloids  to  pass  over  into  less  dispersed 
and  stable  systems  (see  L.  Wöhler's*  observation  on  the  aging  of  col- 
loidal molybdic  and  tungstic  acid).  Several  years  ago  H.  Bechhold 
and  J.  Ziegleri  sought  to  prepare  for  therapeutic  purposes,  with 
the  aid  of  new  and  especially  suitable  protective  colloids,  solutions 
of  such  organic  substances  as  are  insoluble  in  water  (iodoform,  iodo- 
chloroxychinolin,  camphor,  etc.).  They  succeeded  in  thus  preparing 
the  substances,  which,  however,  kept  only  a  few  weeks,  when  they 
would  separate  out  in  crystals.  Obviously  these  substances  are  not 
sufficiently  insoluble  and  they  exhibited  the  adsorption  phenomenon  de- 
scribed on  page  18.  P.  P.  von  Weimarn  made  analogous  observations 
on  the  sol  of  barium  sulphate  in  which  crystals  appeared  at  the  end  of 
six  months.  The  inequality  of  the  particles,  or  more  correctly  the 
''specific  surface,"  obviously  militates  against  the  stability  of  such 
colloid  solutions.  In  the  majority  of  cases,  it  soon  leads  to  the 
"death"  of  the  colloidal  system. 

Furthermore,  we  must  emphasize  that  the  changes  in  the  col- 
loidal system  need  not  always  consist  in  a  diminution  of  the  disper- 
sion. Occasionally  we  find  that  the  particles  become  smaller  with 
the  lapse  of  time,  but  this  has  hitherto  been  observed  only  in  the 
case  of  hydrophile  colloids  (glycogen,  benzopurpurin,  hemoglobin, 
lecithin,  etc.)  (W.  Biltz  and  L.  Gatin-Gruszewska,*  Lemanissier,* 

E.  RÄHLMANN,*!  R.  Zsi GMÜND Y*2). 

Under  some  circumstances  even  electrolytes  may  act  disruptively. 
Thus  B.  G.  Moore  and  H.  E.  Roaf*  observed  that  minutest  traces 
of  electrolytes  are  absolutely  necessary  for  the  stability  of  an  albumin 
solution,  as  was  frequently  pointed  out  by  E.  Jordis  for  hydrophile 
sols.  W.  Biltz  and  H,  von  Vegesack*  observed,  however,  that  in 
the  case  of  dye  solutions,  merely  with  the  lapse  of  time,  marked  in- 
crease in  viscosity  occurs. 

It  may  be  pointed  out  in  connection  with  the  aging  of  jellies,  that 

^  As  yet  unpublished. 


74  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

freshly  poured  gelatin  cylinders  reach  a  practically  constant  modulus 
of  elasticity  at  the  end  of  three  to  four  hours.  This  accords  with  the 
fact  observed  by  F.  Stoffel,  that  crystalloids  diffuse  more  rapidly 
in  quickly  chilled  than  in  slowly  chilled  gelatin,  and  that  this  differ- 
ence disappears  after  several  days  (see  p.  54). 

At  the  outset  we  spoke  of  the  "life  curve  of  colloids,"  of  "aging 
phenomena,"  "death,"  "individual  properties,"  etc.,  and  it  might 
appear  that  these  are  only  similes  borrowed  from  the  organized 
world.  In  my  opinion  the  relationship  is  closer,  and  I  believe  that 
we  may  obtain  a  more  profound  understanding  of  the  phenomena  of 
Life  (so  unintelligible  to  us)  by  a  study  of  such  phenomena  in  colloids. 

Aging  has  hitherto  been  considered,  for  the  most  part,  a  purely 
biological  phenomenon.  In  my  opinion,  we  may  attack  the  problems 
with  the  methods  of  exact  science,  if  we  could  but  separate  two 
groups:  the  organs  (cell  groups),  which  constantly  renew  themselves, 
from  those  which  are  lasting.  We  would,  a  priori,  expect  changes 
in  the  latter  similar  to  those  observed  as  aging  phenomena  in  col- 
loids. We  saw  that  a  rapidly  chilled  gelatin  was  at  first  easily  pene- 
trable for  crystalloids,  but  that  with  time  its  resistance  increased. 
We  may,  therefore,  assume  that  in  young  organs  (fresh  membranes) 
the  exchange  of  matter  by  diffusion  proceeds  more  rapidly.  The 
decrease  in  elasticity,  one  of  the  most  characteristic  phenomena  of 
aging,  may  be  measurably  followed  in  aging  gelatin.  In  fact  it  has 
been  shown  that  for  the  vital  staining  of  nerves  with  methylene  blue, 
young  animals  are  more  suitable  than  old  ones.  With  aging  there 
occurs  shrinking,  which  begins  already  in  intrauterine  life.  In  the 
third  month  of  human  fetal  life  the  water  content  is  94  per  cent,  at 
birth  it  is  69  to  66  per  cent,  in  adult  life  58  per  cent.  We  may  say 
in  general  that  with  aging  there  is  a  decrease  in  the  swelling  capacity 
of  the  organ  colloids.  This  holds  both  for  animal  organisms,  which 
lose  water  as  they  grow  older,  and  for  plants  (dry  leaves  —  lignifi- 
cation). 


Tyndall  Phenomenon.     (From  Wo.  Ostwald.) 


J 


(a)  (b) 

Suspensions  of  Lamp-black, 
(a),  uncoagulated;  (b),  coagulated.     (From  E.  E.  Free.) 


PLATE  I. 


CHAPTER  VI. 

OPTICAL  AND  ELECTRICAL  PROPERTIES  OF  COLLOIDS. 
Optical  Properties. 

Colloidal  solutions,  e.g.,  albumin,  always  show  a  slight  turbidity. 
If  a  strong  ray  of  light  is  passed  through  such  a  solution,  its  path 
may  be  seen  as  a  bright  band.  (See  Plate  I.)  The  "Tyndall  phe- 
nomenon," as  it  is  known,  is  much  more  distinct  if  a  ray  of  light  is 
passed  through  smoke  or  through  a  turbid  suspension,  in  which  case, 
the  reflected  hght  is  polarized.  This  phenomenon  manifests  itself 
if  a  sunbeam  passes  into  a  dark  room.  The  illuminated  dust  par- 
ticles (motes)  appear  bright  against  the  dark  background. 

Michael  Faraday  observed  the  phenomenon  in  the  case  of  gold 
hydrosols  and  he  was  led  to  the  opinion  that  such  solutions,  which  we 
nowadays  call  colloids,  were  nothing  but  extremely  finely  divided 
suspensions  (dispersed  systems).  It  was  a  great  service  to  science, 
when  H.  Siedentopf  and  R.  Zsigmondy  recognized  the  importance 
of  this  phenomenon  for  the  investigation  of  colloids,  and  constructed 
an  instrument  adapted  to  this  purpose  by  passing  the  reflected  light 
into  a  microscope.  In  this  way  they  obtained  bright  pictures  of  the 
suspended  particles  on  a  dark  ground.  Since  neither  of  these  in- 
vestigators paid  any  attention  to  the  representation  of  the  shape  of 
the  particles,  and  devoted  their  attention  only  to  the  reflection  of  a 
point  of  light,  it  was  possible,  by  utiHzing  the  strongest  sources  of 
light  (sun,  arc  lamp),  to  perceive  particles  lying  below  the  limit  of 
microscopic  visibility.  They,  therefore,  called  the  apparatus  the 
ultramicroscope . 

The  great  number  of  fundamental  observations  with  the  ultra- 
microscope  which  we  owe  to  R.  Zsigmondy  *2  and  his  followers  have 
been  repeatedly  mentioned.  R.  Zsigmondy  called  the  particles 
which  he  could  definitely  distinguish  against  the  dark  field,  but 
which  were  far  below  the  limit  of  microscopic  visibility,  submicrons 
(from  6  to  250  fxix).  If  only  a  faint  cone  of  light  could  be  seen,  it 
is  to  be  assumed  in  many  cases,  that  the  smallness  of  the  individ- 
ual particles  precludes  the  recognition  of  each  one.  Such  particles 
(under  6  mm)  be  called  amicrons. 

Most  inorganic  hydrosols,  especially  metals,  form  characteristi- 
cally colored  solutions,  e.g.,  silver  hydrosols  are  brown,  platinum 

75 


76  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

hydrosols  are  greenish  brown,  gold  hydrosols  are  red  but  become  blue 
and  finally  brown  when  electrolytes  are  added.  In  the  ultramicro- 
scope  the  individual  particles  are  not  of  a  uniform  color.  For  in- 
stance, coUargol  has  blue,  red,  violet  and  green  particles,  the  particles 
of  a  red  gold  solution  are  chiefly  green,  those  of  blue  solutions  range 
from  yellow  to  reddish  brown. 

Theoretically,  submicrons  of  the  same  size  should  have  the  same 
color  so  that  the  variety  of  color  in  the  ultramicroscope  indicates 
variation  in  the  size  of  the  particles.  As  a  matter  of  fact,  as  we  have 
said,  the  smaller  submicrons  of  finely  divided  red  gold  hydrosol  are 
almost  all  green  though  there  are  very  small  brown  submicrons. 
There  is  no  entirely  acceptable  explanation  for  the  color  variation  of 
submicrons  of  identical  size. 

The  number  of  particles  visible  in  the  ultramicroscope  is,  in  the 
case  of  hydrophile  colloids,  usually  far  less  numerous  than  might 
have  been  expected  from  their  other  properties.  This  is  the  result 
of  their  inferior  reflecting  power.  If  a  piece  of  swollen  gelatin  is 
immersed  in  water  it  becomes  invisible,  because  no  light  is  reflected 
to  the  eye.  On  this  account  the  ultramicroscope  is  not  suitable  for 
determining  the  number  or  size  of  the  particles  of  hydrophile 
colloids. 

We  may  here  recall  an  observation  of  G.  Quincke*^  which  is 
perhaps  destined  to  be  of  great  importance  for  many  biological 
questions  but  which  deserves  attention,  even  from  a  purely  physical 
standpoint.  G.  Quincke  observed  that  in  the  induced  clarification 
of  mastic,  gamboge,  kaolin  and  India  ink  suspensions,  the  flocks 
usually  separated  on  the  dark  side;  in  spontaneous  clarification  of 
kaolin  turbidity,  however,  they  settled  on  the  light  side.  A  turbid 
solution  of  tannate  of  glue  settled  out,  mostly  on  the  side  towards 
the  light.  He  described  this  phenomenon  as  positive  and  negative 
photodromy.  This  fact  is  suggestive  of  many  of  the  phenomena 
which  H.  SiEDENTOPF*^  observed  in  his  light  reactions  in  the  ultra- 
microscope. 

Jellies,  especially  those  of  higher  concentration,  on  deformation 
by  compression  or  traction,  show  double  refraction  (Kundt).  Nega- 
tive refraction  was  observed  in  the  case  of  gum  arable,  collodion  and 
gelatin,  positive  in  the  case  of  tragacanth  and  cherry  gum.  The 
same  kinds  of  jellies  when  dried  showed  respectively  the  same  kinds 
of  refraction.  If  gelatin  poor  in  water  is  brought  in  contact  with 
gelatin  rich  in  water,  so  that  there  is  a  mutual  interchange  of  water, 
both  become  doubly  refractive.  (M.  W.  Beijerinck.)  On  re- 
peated swelling  and  shrinking  of  jellies,  the  positive  double  refraction 
passes  through  an  isotropic  condition  into  a  negative.     (Quincke.) 


OPTICAL  AND  ELECTRICAL  PROPERTIES  OF  COLLOIDS      77 

Chlorids  and  nitrates  diminish  the  double  refraction;    sulphates  are 
without  effect.     Phenols  change  the  direction  of  the  refraction. 

The  phenomenon  is  important  for  the  understanding  of  the  double 
refraction  of  organized  structures  (plant  fibers,  muscle,  horn,  etc.). 

Electrical  Properties. 

If  two  electrodes  are  placed  in  a  solution  of  a  hydrosol  as  free  as 
possible  of  electrolytes,  and  a  current  is  allowed  to  pass  through,  we 
immediately  notice  the  movement  of  the  colloid  to  one  of  the  elec- 
trodes. Various  suspensions  (suspensions  of  clay,  rosin,  etc.),  as 
well  as  most  hydrophobe  hydrosols  migrate  to  the  anode,  whereas 
the  colloidal  metal  hydroxids  (iron  or  aluminium  oxid  hydrosol,  etc.) 
move  to  the  cathode.  Hydrophile  colloids  (albumin,  etc.)  exhibit, 
if  almost  free  from  electrolytes,  no  definite  recognizable  directive 
tendency.  The  zone  of  H  ion  concentration  in  which  there  is  no 
migration  has  been  named  by  L.  Michaelis  the  isoelectric  zone. 
The  addition  of  acids  causes  migration  of  these  colloids  to  the 
cathode,  alkalies  to  the  anode;  they  then  behave  as  if  they  were 
salts  of  the  acids  or  alkalies  involved,  and  this  may  actually  corre- 
spond with  the  facts  (see  p.  149  et  seq.). 

This  movement  of  suspensions  and  hydrosols  against  the  water, 
under  the  influence  of  the  electric  current,  is  called  cataphoresis. 

The  colloid  particles  behave  like  ions  and  their  speed  of  migration 
is  similar  in  rate.  Zsigmondy  calculated  from  the  speed  of  migration 
(0.002  mm.)  and  the  diameter  (50  mm)  of  the  particles  of  a  colloidal 
silver  solution,  that  the  particles  were  charged  with  297  X  10"^" 
electrostatic  units  which  is  the  equivalent  of  62  elemental  units. 
Such  a  particle  is  in  a  certain  sense  an  ion  of  62  valencies. 

If  a  protective  colloid  (albumin,  gelatin,  etc.)  is  added  to  a  sus- 
pension, the  latter  acts  as  if  its  entire  mass  was  composed  of  the 
protective  colloid.  The  commercial  inorganic  colloids  (collargol, 
lysargin,  etc.)  do  not  behave  in  an  electric  current  as  does  pure 
colloidal  silver,  but  as  albumins  or  albumoses.  Their  direction  may 
be  changed  at  will  by  the  addition  of  acids  or  alkalis. 

The  process  may  be  reversed,  that  is,  the  water  may  be  moved 
under  the  influence  of  electrical  difference  in  potential,  provided 
the  suspension  is  held  fixed.  The  experimental  procedure  is  as 
follows:  Instead  of  a  clay  suspension  we  choose  a  porous  clay  wall 
D  (Fig.  8),  permeable  for  water,  by  means  of  which  a  U-shaped 
tube  is  divided  into  two  parts.  If  the  tube  is  filled  with  water  and 
into  each  branch  an  electrode  is  introduced,  the  water  moves  under 
the  influence  of  the  electric  current  and,  in  fact,  it  will  rise  on  the 
cathode  side  until  it  exerts  a  certain  pressure  against  the  anode 


78 


COLLOIDS  IN  BIOLOGY  AND   MEDICINE 


side.  This  process  is  called  electro-endosmosis.  In  principle  the  two 
phenomena  are  the  same.  For  the  exact  study  of  electrical  migra- 
tion, which  was  first  investigated  by  G.  Wiedemann  and  G.  Quincke, 
electro-endosmosis  proved  to  be  experimentally  more  easily  avail- 
able.   A.  CoEHN*  has  shown  that  it  is  a  general  and  quantitative  law, 


V 


^S 


-^-^.   jy 


Fig.  8.     Apparatus  for  electro-endosmosis.    (H.  Freundlich.) 

that  substances  possessing  higher  dielectric  constants  are  positively 
charged  when  brought  into  contact  with  substances  of  lower  dielectric 
constants. 

We  have  already  seen  that  numerous  substances  are  negatively 
charged  in  respect  to  water  while  others  are  positively  charged  and 
that  under  the  influence  of  the  electric  current  there  occurs  a  move- 
ment of  the  suspended  substance  or  of  the  water.  This  migration 
may  be  influenced  by  the  addition  of  electrolytes.  J.  Perrin  used 
diaphragms  of  porous  carborundum  and  of  naphthalin  and  measured 
the  amount  of  water  which  passed  through  the  porous  wall  by  en- 
dosmosis,  under  the  influence  of  an  electric  current. 

In  the  following  table  +  indicates  passage  to  the  positive,  —  to 
the  negative  pole. 


Diaphragm. 

Solution. 

Amount  of  fluid 

transferred  in  cmm. 

per  minute. 

Carborundum 

Carborundum 

Carborundum 

Carborundum 

Carborundima 

Carborundum 

Naphthalin  ^ 

0.02      mol.  HCl 
0.008    mol.  HCl 
0.002    mol.  HCl 
dist.  water 
0.0002  mol.  KOH 
0.002    mol.  KOH 
0.02      mol.  HCl 
0.001    mol.  HCl 
0.0002  mol.  HCl 
0.0002  mol.  KOH 
0.001    mol.  KOH 

+  10 
0 

-  15 

-  50 

-  60 
-105 
+  38 

Naphthalin 

Naphthalin 

+  28 
+    3 

Naphthalin.  .           

-  29 

Naphthalin .                         

-  60 

I  I  tried  to  cause  naphthalin  suspensions  to  migrate  in  strong  currents  (400  volts)  but  could  not 
observe  any  cataphoresis  either  in  neutral,  in  acid  or  in  alkaline  solution.  Nor  could  I  obtain  a 
migration  of  suspensions  of  naphthol  and  naphthylamin  in  neutral,  very  faintly  acid  or  very  faintly 
alkaline  solution  (hitherto  unpublished). 


OPTICAL  AND  ELECTRICAL  PROPERTIES  OF  COLLOIDS      79 

It  follows  from  this  that  the  negative  charge  of  negative  diaphragms 
(as  is  evident  from  the  table)  increased  in  alkaline  solution.  With 
a  decrease  in  the  OH  ion  concentration  (with  naphthalinj  or  the  in- 
crease in  the  H  ion  concentration  (with  carborundum)  a  point  was 
reached  in  which  there  was  no  difference  in  potential  between  w^ater 
and  diaphragm.  With  further  increase  in  the  H  ion  concentration 
the  diaphragm  took  on  a  stronger  positive  charge. 

If  a  positive  diaphragm  is  chosen,  as  for  instance  chromium  chlorid, 
the  conditions  are  reversed. 

J.  Perrin  studied  the  influence  of  salts  in  the  presence  of  acids 
and  bases.  It  was  sho^^^l  that  they  cause  a  loss  of  charge  with  moder- 
ate concentrations  and,  in  fact,  that  the  strength  of  their  action  de- 
pended, in  the  case  of  positively  charged  diaphragms,  upon  the 
valence  of  the  anions,  while  in  the  case  of  negatively  charged  dia- 
phragms it  depended  upon  the  valence  of  the  cations.  With  higher 
concentration  of  polj^^alent  anions  or  cations  a  loss  of  charge  may 
occur. 

Upon  calculating  the  concentration  of  salt  which  just  halves  the 
amount  of  fluid  (r  in  mm./minutes)  transferred,  the  follo'^ing  figures 
were  obtained: 


Diaphragm. 

Charge. 

Salt. 

V 

1  =  1 

(NaBrorKBr). 

Carborundum 

XaBr 

50 

1 

Carborundum 

— 

Ba(X03)2 

2 

25 

Carborundum 

— 

La(X03)3 

0.1 

500 

Chromium  chlorid 

+ 

KBr 

60 

1 

Chromium  chlorid 

+ 

]MgS04 

1 

60 

Chromium  chlorid 

+ 

K3Fe(CX)6 

0.1 

600 

The  last  column  -  shows  us  that  the  discharging  effect  increases 

V 

with  the  valence  of  the  anions  as  1  :  25  :  500  and  in  the  case  of 
cations  as  1  :  60  :  600.  In  the  section  on  "  flocculation, "  we  shall 
see   a  vers"  remarkable  application  of  this  phenomenon. 

Of  the  many  theories  proposed  to  explain  these  circumstances, 
that  of  H.  Freundlich  and  L.  ]\Iichaelis,  according  to  which  the 
various  ions  are  adsorbed  to  different  degrees,  seems  to  us  most  prob- 
ably correct;  in  point  of  fact,  the  H  and  OH  ion  have  especially 
high  adsorption  coefficients.  Since  a  separation  of  anion  and  cation 
usually  cannot  occur,  there  arises  a  difference  in  potential  at  the 
surface  between  the  dispersed  phase  and  the  water.  An  indifferent 
substance  in  a  weakly  acid  solution  will  adsorb  H  ions  and  become 


80  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

positively  charged  in  respect  to  the  fluid;  in  an  alkaline  fluid  it  will 
become  negatively  charged.  If  the  dispersed  phase  itself  has  basic 
or  acid  properties  it  will  behave  in  pure  water  like  a  cation  or  an 
anion  respectively.  If  to  an  acid  suspension,  e.g.,  clay,  an  alkali  is 
added,  K  ions  are  adsorbed,  OH  ions  are  concentrated  and  held  at 
the  outer  film  and  the  negative  charge  is  thereby  increased.  The 
reverse  occurs  upon  adding  acids;  the  charge  is  released  and  may 
even  take  the  opposite  sign.  According  to  this  view,  colloids  wan- 
dering to  the  anode  are  discharged  only  by  cations;  those  travelling 
to  the  cathode  only  by  anions.  We  have  seen  that  polyvalent  ions 
have  a  considerably  greater  discharging  power,  which  increases  with 
their  valence.  This  experimental  fact  accords,  as  do  all  the  others, 
with  the  theory  propounded. 

In  the  case  of  hydrophile  colloids,  it  suffices  for  us  to  assume  that 
we  are  dealing  with  very  large  amphoteric  molecules  which  become 
cations  in  acid  solution  and  anions  in  alkaline  solution. 


Salting  Out. 

If  large  quantities  of  a  neutral  salt,  for  instance  ammonium 
sulphate,  are  added  to  a  solution  of  a  hydrophile  colloid  (albumin, 
globulin,  casein,  albumose,  silver  protected  by  a  protective  colloid, 
etc.),  or  certain  inorganic  hydrosols,  e.g.,  sulphur,  the  colloid  is  thrown 
out,  but  it  redissolves  upon  dilution  with  water.  This  is  the  process 
of  salting  out,  as  practised  technically.  If,  for  instance,  enough  com- 
mon salt  is  added  to  an  aqueous  solution  of  phenol,  the  phenol 
separates  out.  We  might  regard  this  as  a  withdrawal  of  water  by 
means  of  the  electrolyte,  since  we  know  from  the  observations  of 
recent  years  that  ions  form  hydrates  in  aqueous  solution,  i.e.,  every 
ion  attracts  a  greater  or  smaller  number  of  molecules  of  water.  I 
have,  however,  been  unable  to  discover  any  relation  between  the 
salting  out  process,  from  the  figures  for  the  hydration  of  a  number 
of  different  ions  as  calculated  by  E.  H.  Riesenfeld  and  B. 
Reinhold*. 

The  more  closely  a  hydrophile  colloid  approaches  the  crystalloid 
condition,  the  greater  is  the  concentration  of  salt  required  for  salting 
out.  Thus,  for  instance,  the  alhumoses  are  classified  in  accordance 
with  concentration  of  salt  required  for  their  precipitation  (see 
p.  166.) 

As  early  as  1907,  Bechhold  had  already  called  attention  to  the 
relation  between  salting  out,  and  particle  size  and  salt  concentration. 
S.  Oden  produced  the  experimental  evidence  that  reversible  sul- 
phur and  silver  hydrosols  could  actually  be  separated  in  fractions, 


OPTICAL   AND  ELECTRICAL   PROPERTIES  OF   COLLOIDS      81 

distinguishable  by  the  size  of  their  particles,  Ijy  "fractional  coagula- 
tion/' ^  i.e.,  by  the  addition  of  salt  solutions  of  progressively  increas- 
ing strength. 

The  influence  of  neutral  salts  exhibits  regularities  which  are  of 
great  importance  in  a  number  of  biological  phenomena  and  which  we 
shall  repeatedly  encounter.  The  salting  out  of  hydrophile  colloids, 
gelatinization,  the  irritability  of  muscle  and  nerve,  the  permeability 
of  cell  membranes  (blood  corpuscles,  etc.),  the  swelling  of  membranes 
and  many  other  phenomena  are  thus  related  to  a  group  of  physico- 
chemical  properties  of  solutions,  whose  relationship  is  indubitable 
even  though  the  true  basis  is  not  clear.  With  H.  Freundlich  we 
shall  describe  these  effects  of  neutral  salts  as  lyotropic  (solution 
changing),  and  we  shall  study  them  more  closely. 

Most  inorganic  salts  increase  the  surface  tension  of  water  and  from 
a  table  of  W.  K.  Röntgen  and  J.  Schneider  we  obtain  the  following 
series  for  the  increase  in  the  surface  tension  by  the  alkaline  iodids: 

Na  >  K  >  Li  >  NH4. 

With  the  anion  of  various  alkalis: 

CO3  >  SO4  >  CI  >  NO3  >  I. 

Though  the  compressibility,  the  solubility  and  the  viscosity  of 
water  are  changed  in  a  similar  order  by  neutral  salts,  the  relation- 
ship is  much  more  fundamental.  Neutral  salts  may  accelerate  or 
impede  catalytic  effects,  such  as  the  inversion  of  cane  sugar,  the 
saponification  of  esters  and  the  changing  of  acetone  into  diacetone 
alcohol.  The  action  in  acid  solutions  is  usually  the  reverse  of  what 
it  is  in  alkaline  solutions  as  has  been  shown  by  R.  Höber.  In  acid 
media  the  acceleration  by  cations  is 

Li  >  Na  >  K  >  Rb  >  Cs, 
in  alkaline  media 

Cs  >  Rb  >  K  >  Na  >  Li. 

For  anions  in  acid  solutions  the  following  order  holds: 
I  >  NO3  >  Br  >  CI  >  CH3CO2  >  SO4. 

In  alkaline  solution  the  series  is  reversed. 

In  neutral  solution  also  the  lyotropic  series  holds,  although  small 
changes  in  the  arrangement  may  exist  for  some  of  the  ions. 

^  Since  reversible  hydrosols  are  considered  I  might  describe  the  procedure 
preferably  as  "fractional  salting  out." 

It  is  particularly  interesting  to  know;  that  according  to  Oden  and  Oholm  the 
particles  do  not  coalesce  but  retain  their  identity  and  when  they  are  redissolved, 
there  are  as  many  particles  in  solution  as  before  the  salting  out. 


82  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

We  encounter  such  lyotropic  series  regularly  in  the  salting  out  of 
hydrophil  ecolloids  (see  Albumin,  p.  146  et  seq.,  Lecithin,  p.  140, 
Gelatin,  p.  161)  and  in  many  biological  phenomena,  where  they  tend 
to  cause  either  a  precipitation  or  a  concentration. 

However,  we  must  not  assume  that  the  action  is  such  that  in  one 
case  the  anion  alone  is  active,  and  in  the  other  the  cation  alone. 
There  are  many  reasons  for  believing  that  there  is  an  antagonism 
between  anions  and  cations,  and  that  the  action  of  the  cation  is  more 
or  less  powerfully  increased  or  diminished  depending  upon  the 
anions  present.  If  in  a  given  instance  we  speak  of  the  cations 
causing  precipitation  or  dehydration,  we  always  mean  the  difference 
between  the  effect  of  the  cation  and  the  opposed  action  of  the  anion 
present,  in  which,  however,  the  action  of  the  cation  predominates. 

The  divalent  cations  Mg,  Ca,  Sr  and  Ba  act  more  strongly  as  pre- 
cipitants  than  the  monovalent  cations.  Biologically  they  are  im- 
portant in  connection  with  alkali  salts,  inasmuch  as  small  quantities 
of  calcium  salts  are  able  to  replace  considerably  larger  quantities  of 
alkali  salts,  e.g.,  Na  or  K.  This  greater  effect  may  even  lead  to  an 
antagonism  between  the  two;  it  has  been  thoroughly  studied  by 
Wo.  Pauli  and  H.  Handovsky.*^  It  is  sufficient  to  mention 
that  in  the  case  of  alkali  albumin,  Ca  salts  form  a  less  ionized 
Ca  albumin  combination  from  a  more  ionized  Na  albumin  combi- 
nation. According  to  the  law  of  mass  action  small  quantities  of  Ca 
may  be  replaced  by  larger  quantities  of  Na.  We  may  thus  under- 
stand the  significance  of  the  Ca  content  in  all  physiological  fluids. 
Mg,  Sr  and  Ba  act  in  a  similar  manner.  They  have  in  addition 
certain  specific  properties  which  obscure  the  relations. 

Large  quantities  of  alkaline  earths  cause  irreversible  changes  in 
many  biocolloids,  that  is,  they  produce  insoluble  compounds  with 
them.  Electrolytes  may  exert  another  effect  on  the  biocolloids; 
they  may  cause  "flocculation,"  a  phenomenon  we  shall  now  study 
more  closely. 

Flocculation. 

If  albumin  is  boiled,  it  coagulates.  A  coagulation  of  albumin  may 
also  be  produced  by  the  addition  of  ammonium  sulphate.  Though 
the  latter  process  may  be  reversed  by  dilution  with  water,  the  boiled 
albumin  cannot  again  be  brought  to  its  fluid  condition,  by  any 
physical  means.  The  hydrophile  colloid  has  been  converted  into  a 
hydrophobe  colloid.  For  uniformity's  sake,  we  shall  consider  as  co- 
agulations only  such  processes  as  cause  an  irreversible  change. 

If  we  heat  a  very  dilute  albumin  solution,  there  is  apparently  no 
coagulation;    at   most   the   fluid   becomes   slightly   opalescent.     In 


OPTICAL  AND  ELECTRICAL   PROPERTIES  OF   COLLOIDS      83 

reality  a  coagulation  has  occurred  here,  since  the  addition  of  a  drop 
of  acetic  acid  or  a  little  ammonium  sulphate  produces  flocks.  The 
process  is  therefore  called  flocculation.  These  albumin  flocks  are  in- 
soluble in  water. 

Only  by  the  second  process,  by  the  agglomeration  of  the  smallest 
particles  (see  Plate  I,  h  and  c)  under  the  influence  of  the  acetic  acid 
or  ammonium  sulphate  was  it  possible  to  separate  the  two  phases 
(water/albumin).  This  occurs  in  a  way  similar  to  that  in  which 
the  drops  of  rain  are  formed  under  certain  conditions  by  the  union 
of  particles  of  mist.  The  union  is  preceded  by  a  slowing  of  the 
Brownian-Zsigmondy  movement,  as  has  been  shown  with  the  ultra- 
microscope  by  V.  Henri.* 

Flocculation  is  an  electrical  'phenomenon.  It  is  brought  about  by 
electrolytes  as  well  as  hy  colloids  of  opposite  electric  charge,  as  well  as 
by  ultraviolet  and  Roentgen  rays  (the  action  of  rays  is  much  weaker 
than  the  action  of  electrolytes).^  If  we  shake  purified  lampblack 
with  water  we  get  a  suspension  which  remains  turbid  for  weeks. 
If  we  pour  a  few  drops  of  an  alcoholic  mastic  solution  into  water, 
it  remains  milky  for  weeks  and  even  years.  We  have  seen  that  the 
hydrophobe  inorganic  hydrosols,  such  as  colloidal  arsenic  sulphid, 
platinum  sol  prepared  according  to  Bredig's  method,  gold  sol  pre- 
pared according  to  R.  Zsigmondy's  method,  are  permanent  for 
months  provided  the  solution  is  free  from  electrolytes.  Addition 
of  electrolytes  causes  an  irreversible  flocculation  of  these  hydrophobe 
colloids. 

The  process  is  to  be  sharply  distinguished  from  salting  out,  i.e.,  the 
reversible  precipitation  of  albumin,  albumoses,  etc.,  from  solution  by 
large  quantities  of  salt. 

The  phenomenon  of  flocculation  is  encountered  especially  in  the 
precipitation  reactions  of  albumins  and  in  Chapter  XIII  on  Im- 
munity Reactions,  where  it  plays  an  important  role  in  the  agglutina- 
tion of  bacteria  by  precipitins.  Moreover  the  precipitation  of  gold 
hydrosols  by  cerebrospinal  fluid  has  acquired  great  significance  in 
the  diagnosis  of  mental  diseases  (see  p.  354). 

To  cause  flocculation,  a  certain  minimum  amount  of  electrolyte  as 
as  well  as  of  the  dispersed  phase  is  required.  Below  these  limits, 
which  are  characteristic  for  every  electrolyte,  no  flocculation  occurs 
even  after  months.  H.  Bechhold  called  this  minimum,  the  "elec- 
trolyte threshold"  and  the  "suspension  threshold.'' 

The  rate  of  flocculation  is  dependent  on  the  concentration  of  the 
suspension  and  of  the  electrolytes,  i.e.,  the  more  concentrated  the  sus- 

"■  Specific  immunity  reactions,  in  which  precipitations  occur,  are  probably  not 
electrical  (see  p.  197). 


84  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

pension  and  the  electrolytes  within  certain  limits,  the  greater  is  the 
rapidity  of  flocculation.  The  dependence  upon  the  concentration  of 
the  electrolytes  is  especially  noticeable  in  the  vicinity  of  the  elec- 
trolyte threshold.  Further  away  from  the  electrolyte  threshold  the 
rapidity  of  flocculation  is  less  dependent  upon  the  concentration  of 
the  electrolyte.     (H.  Bechhold.*^) 

Under  certain  conditions,  an  excess  of  electrolytes  may  lead  to 
re-solution.  This  phenomenon  is  called  peptisation.  Graham  first 
observed  this  occurrence  upon  treating  ferric  oxid  gel  with  ferric 
chlorid  and  compared  it  to  the  formation  of  water-soluble  peptone 
from  coagulated  albumin  and  hydrochloric  acid.  As  a  matter  of 
fact,  peptisation  probably  depends  upon  a  renewal  of  electric  charge 
by  the  excess  of  electrolytes. 

As  has  been  shown  by  H.  Freundlich  and  his  pupils  Ishizaka 
and  ScHUCHT  *^,  these  facts  furnish  an  indication  that  the  more  sen- 
sitive a  substance  is  to  flocculation,  the  more  will  it  be  adsorbed. 

The  great  majority  of  colloids  migrate  to  the  anode,  and  on  floccu- 
lation the  cation  is  of  much  the  greatest  importance;  the  anion  plays 
but  a  subordinate  role.  The  conditions  are  reversed  in  the  case  of 
the  few  colloids  which  migrate  to  the  cathode.  However,  R.  Burton 
has  shown  that  with  increase  of  electrolyte  concentration  the  rate  of 
migration  of  the  colloid  becomes  constantly  diminished  and  finally 
the  direction  may  change.  At  the  stage  when  the  concentration  of 
electrolytes  is  such  that  reversal  occurs,  in  other  words  the  isoelectric 
zone,  flocculation  is  most  rapid.  The  increase  in  the  action  of  the 
cations  is  out  of  proportion  to  the  increase  in  their  valence.  The 
electrolyte  concentration  necessary  for  the  flocculation  of  a  mastic 
suspension  is  FeCls  :  BaCl2  :  NaCl  =  1  :  50  :  1000.  There  are  also 
certain  anomalies  (see  H.  Bechhold,*i  as  well  as  M.  Neisser  and 
U.  Friedemann*)  in  connection  with  the  rate  of  migration  of  the 
ions,  as  well  as  in  the  electrolytic  dissociation  and  especially  in  the 
ionization  pressure  of  electrolytes.  In  general  the  above  relation 
between  the  action  of  the  cations  is  maintained.  The  powerful 
flocculating  action  of  the  H  ion  without  doubt  depends  upon  its 
high  speed  of  migration  while  the  OH  ion  has  a  corresponding  action 
upon  colloids  which  migrate  to  the  anode. 

Flocculation  by  means  of  trivalent  iron  and  aluminium  salts  show 
peculiar  anomalies.  These  were  discovered  by  M.  Neisser  and 
U.  Friedemann*  and  also  by  H.  Bechhold*^  and  called  irregular  series. 
Later  these  phenomena  of  Inhibition  Zones  were  followed  further 
by  0.  Teague  and  B.  H.  Buxton*^  and  also  by  A.  Lottermoser.* 
An  example  from  my  published  paper  will  best  explain  the  phenom- 
enon.    XXX  means  strong  flocculation,   X  X  medium,  X  none. 


OPTICAL  AND  ELECTRICAL  PROPERTIES  OF  COLLOIDS     85 


Concentration 

of  mastic 

0.01 

0.005 

0.0C25 

0.001 

0.0005 

0.00025 

solution. 

1/2 

XXX 

XXX 

XXX 

XX 

XXX 

XX 

1/4 

.XXX 

XXX 

XX 

XX 

XXX 

0 

1/8 

XXX 

XXX 

0 

0 

XXX 

X 

1/16 

XXX 

XX 

0 

0 

XX 

XXX 

1/32 

XXX 

0 

0 

0 

0 

XXX 

Mol. 

Al2(SO^)3 


The  discoverers  explain  the  phenomenon  by  saying  that  the  salts 
mentioned  undergo  strong  hydrolytic  dissociation  so  that  the  col- 
loidal iron  and  aluminium  hydroxid  present  in  the  solution  act  as 
''protective  colloids"  somewhat  like  gelatin  or  albumin. 

The  flocculation  hitherto  described  occurs  only  with  true  hydro- 
phobe colloids.  If  the  individual  particles  possess  a  film  of  native 
albumin,  gelatin,  an  albumose  (such  as  Na  lysalbinate),  dextrin,  etc., 
the  particles  act  as  if  they  were  the  "protective  colloids"  involved 
(see  p.  77);  in  other  words  they  are  salted  out  only  by  very  large 
amounts  of  electrolytes.  The  colloid  precipitate  can  be  redissolved 
in  water  providing  the  protective  colloid  is  not  denatured  by  the 
electrolytes.  On  this  account  all  the  colloidal  metal  sols  used  thera- 
peutically such  as  collargol,  bismon,  etc.,  are  "stabilized"  by  hy- 
drophile colloids.  Without  this  protection  it  would  be  impossible 
to  preserve  them  for  any  length  of  time  and  they  would  be  floccu- 
lated when  they  were  prepared  for  intravenous  use  by  dilution  with 
physiological  salt  solution. 

R.  ZsiGMONDY  considers  as  characteristic  of  hydrophile  colloids, 

the  protection  they  give  to  his  gold  hydrosol  against  flocculation  by 

electrolytes.     He  designates  as  the  gold  figure,  the  amount  (in  mg.) 

of  the  colloid  in  question,  which  was  just  sufficient  to  prevent  the 

2 
flocculation  of  10  c.c.  of  a  gold  sol.  by  1  c.c.  -  NaCl  sol.     The  floccu- 

lation  is  accompanied  by  a  change  in  color  from  red  to  blue,  and  it  is, 
sufficient  to  observe  this  change. 

The  following  data  from  R.  Zsigmondy  illustrate  this  point: 


Colloid. 

Gold  figure  in  mg. 

Gelatin 

Casein.   . . 

0.005-0.01 
0  01 

Egg  albumen 

Gum  arable 

0.06-0.3    (depending  upon  its  origin  and  ash  content) 
0.15-0.25 

Tragacanth 

Dextrin 

2± 
10-20 

Potato  starch 

Sodium  stearate .... 
Sodium  stearate .... 
Sodium  oleate 

25 

10          (added  at  60°) 
0 .  001      (added  boiling  hot) 
0.4-1 

86  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Peptones  have  no  protective  action  at  all  [Peptones  may  cause 
precipitation.  E.  Zunz,  Bull.  Soc.  Roy.  des  Sc,  Med.,  et  Nat.,  June 
11,  1906.  Tr.],  whereas  some  of  the  albumoses,  especially  sodium 
lysalbinate  and  sodium  protalbinate  have  a  very  powerful  protective 
action  which  C.  Paal  utilized  in  preparing  a  large  number  of  inor- 
ganic colloids. 

Sensitiveness  to  flocculation  may  vary  considerably  with  the 
nature  of  the  protective  colloid. 

The  flocculation  of  hydrosols  may  be  brought  about  not  only  by 
electrolytes  but  also  by  hydrosols,  providing  they  have  an  opposite 
electric  charge,  as  has  been  shown  by  W.  Biltz.  Thus,  for  instance, 
arsenic  trisulphid,  gold  and  platinum  hydrosols,  etc.,  are  flocculated 
by  ironoxid,  aluminiumoxid,  chromiumoxid  hydrosols,  etc.  A  proper 
relative  mixture  is  required,  which  means  the  charge  of  the  positive 
sol  must  be  counterbalanced  by  the  charge  of  the  negative  sol.  If 
a  sol  is  in  excess,  no  flocculation  occurs,  and  the  entire  complex  con- 
sisting of  both  colloids  migrates,  when  placed  between  two  elec- 
trodes, in  the  direction  of  the  sol  (J.  Billiter*)  which  is  in  excess. 
This  explains  why  protective  colloids  may  under  some  circumstances 
produce  flocculation  instead  of  protection,  namely,  when  they  are 
added  in  minimal  quantities.  Thus,  for  instance,  mastic  emulsions 
are  flocculated  by  0.0003  to  0.0001  per  cent  of  gelatin  (H.  Bechhold 
and  also  M.  Neisser  and  U.  Friedemann).  Hydrochloric  acid  in  a 
dilution  incapable  of  producing  flocculation  by  itself  can  coagulate 
gold  hydrosol,  mastic  or  oil  emulsion  in  the  presence  of  one  part  of 
gelatin  per  million. 

As  has  already  been  mentioned  we  must  not  confuse  the  salting 
out  of  hydrophile  colloids  with  flocculation,  though  there  are  border- 
line cases  which  complicate  the  phenomenon.  If  we  add  for  instance 
a  salt  of  a  heavy  metal  to  a  dilute  albumin  solution,  a  more  or  less 
irreversible  albumin-metal  compound  will  form,  dependent  on  the 
nature  of  the  salt  upon  which  the  excess  of  albumin  acts  as  a  pro- 
tective colloid.  Acids  may  cause  a  loss  of  charge  and  the  metal  salt 
may  then  exhibit  some  flocculating  and  some  salting  out  action. 
Such  a  case  might  occur,  for  instance,  on  adding  zinc  sulphate  to 
albumin  (studied  by  Wo.  Pauli).     The  process  is  as  follows: 

ZnS04  +  Albumin. 

0.05  n maximum  flocculation  (irreversible  till  1  n). 

In precipitate  disappears. 

2  n precipitate  reappears  (reversible) . 

4  n maximum  precipitate  (reversible). 

There  are  transitions  between  hydrophobe  and  hydrophile  col- 
loids which  are  responsible  for  the  transitions  between  flocculation 


OPTICAL  AND  ELECTRICAL  PROPERTIES  OF  COLLOIDS     87 

and  salting  out.  Cholesterin  and  lecithin  may  be  regarded  as  such 
transitional  substances,  whose  flocculation  has  been  thoroughly 
studied  by  0.  Porges  and  E.  Neubauer.*  Cholesterin  closely  ap- 
proaches the  hydrophobe  colloids,  lecithin  the  hydrophile,  and  on 
this  account  the  former  is  irreversibly  precipitated  by  salts  and  the 
latter  reversibly.  Yet  the  concentration  of  alkaline  earths  required 
to  precipitate  lecithin  is  considerably  less  than  for  the  more  hydro- 
phile albumin,  and  conversely  much  greater  concentration  of  salt  is 
required  for  the  flocculation  of  Cholesterin  than  is  the  case  with  true 
hydrophobe  sols.  In  the  case  of  lecithin,  the  "irregular  series" 
occurs  even  with  neutral  salts  (magnesium  and  ammonium  sulphate). 
As  will  be  seen  in  Chapter  XIII,  such  transitions  from  hydrophobe 
suspensions  to  hydrophile  colloids  may  be  artificially  produced  with 
emulsions  of  bacteria. 

Much  has  been  written  on  the  theory  of  salting  out  and  flocculation. 
No  one  theory  accounts  for  all  the  individual  facts,  yet  the  following 
explanation  is  generally  useful.  Flocculation  is  brought  about  by 
the  coming  together  of  small  particles  to  form  larger  complexes. 
These  agglomerations  always  occur  under  the  influence  of  electric 
forces  and  in  fact  the  optimum  for  reversible  salting  out  is  in  the  iso- 
electric zone  (see  p.  158).  The  process,  must,  therefore,  be  brought 
about  by  a  discharge  of  the  particles.  Risdale  Ellis  has  shown  by 
researches  on  oil  emulsions,  that  the  charge  at  the  interface  between 
water  and  the  dispersed  phase  is  probably  reduced  to  a  minimum  by 
the  addition  of  precipitating  electrolytes.  The  smaller  this  charge 
is  at  the  interfaces  (electric  double  layer),  the  more  readily  the 
double  layer  is  broken  down,  resulting  in  a  union  of  the  suspended 
fluid  or  solid  particles. 

In  the  case  of  anodic  colloids,  cations  of  an  electrolyte,  and  in  the 
case  of  cathodic  colloids,  anions  or  an  oppositely  charged  colloid, 
lessen  or  neutralize  the  electric  charge,  so  that  the  particles  may 
unite.  (See  H.  Freundlich's  "  Kapiflarchemie  "  and  Wo.  Ostwald's 
"Grundriss  der  Kolloidchemie.") 

Irreversible  precipitation  of  metal  hydrosols  frequently  occurs  out- 
side the  isoelectric  zone  and  the  electrolyte  threshold  is  not  as  sharp 
as  with  reversible  hydrosols. 

Radioactive  Substances  as  Colloids. 

It  has  been  demonstrated  by  electric  migration  and  dialysis  that 
radioactive  substances  occur  in  colloidal  solution.  In  view  of  the 
great  biological  significance  of  such  substance  we  shall  explain  the 
facts  more  fully. 


88  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

GoDLEWSKi  *  found  upon  electrolysis  of  the  radium  emanations  and 
their  products  that  the  radioactive  substances  migrated  mostly  to 
the  cathode  in  acid  solutions  and  to  the  anode  in  alkaline  solutions. 
Paneth  dialyzed  a  solution  of  radio-lead  nitrate  against  pure  water 
in  a  parchment  thimble  and  greatly  concentrated  the  RaE  and 
polonium  in  the  thimble  but  the  relation  between  RaD  to  lead  was 
not  changed.  Both  phenomena  are  characteristic  of  colloids  and 
indicate  the  colloid  properties  of  radioactive  substances;  this  led 
them  to  continue  their  investigations.  Godlewski  *  successfully 
adsorbed  RaA,  RaB  and  RaC  with  inorganic  colloids  and  concen- 
trated the  radioactive  substances  by  precipitating  the  latter.  He 
was  successful  not  only  with  radium  but  with  actinium,  mesothorium 
and  uranium.  The  concentration  of  radioactive  substances  by 
adsorption  on  colloidal  silicic  acid  (Ebler,  Fellner)  has  attained 
great  practical  value  in  the  manufacture  of  radium  preparations. 
In  fact  RaA  and  RaC  may  be  collected  from  acid  solutions  simply 
on  filter  paper  and  by  burning  the  paper  a  highly  active  ash,  free  from 
RaB,  may  be  obtained.  It  is  evident  that  in  solution  radioactive 
elements  undergo  hydrolysis  with  the  formation  of  a  colloidal  radio- 
hydrosol. 


CHAPTER  VII. 

METHODS    OF   COLLOID   RESEARCH. 

A  FIELD  of  research  extending  as  does  colloid  chemistry  into  so 
many  other  branches  of  science  must  be  served  by  countless  methods. 
Purely  chemical  as  well  as  physical  and  biological  methods  which 
the  investigator  of  colloids  utilizes  in  his  studies,  have  been  so  well 
developed  and  so  thoroughly  described  in  technical  literature,  that  it 
is  needless  to  discuss  them  more  fully  here. 

There  are,  however,  several  methods  which  are  peculiar  to  colloid 
investigation,  and  we  shall  consider  them  here.  Unfortunately,  the 
limits  of  their  usefulness  are  as  yet  unestablished;  for  some  one  to 
establish  them  would  be  highly  desirable. 

In  discussing  the  following  methods,  I  have  not  sought  complete- 
ness but  have  considered  only  those  which  have  proved  practical; 
I  have  personally  tested  most  of  them. 

To  determine  whether  a  solution  of  a  substance  is  colloidal  in 
character  it  must  be  tested  by  dialysis,  ultrafiltration  or  diffusion. 

Dialysis  is  a  purely  qualitative  method  which  determines  whether 
or  not  a  substance  is  colloidal  in  character,  i.e.,  whether  or  not  it 
consists  of  large  particles. 

Ultrafiltration  in  most  cases  may  be  used  instead  of  dialysis;  it 
works  much  more  rapidly  and  above  all  permits,  in  addition,  quan- 
titative experiments,  and  consequently  has  a  much  broader  utility. 

Both  methods  serve  to  separate  colloids  from  crystalloids.  This 
separation  occurs  with  dialysis  if  the  water  surrounding  the  dialyzer 
is  frequently  renewed;  it  occurs  with  ultrafiltration,  provided  the 
substance  on  the  funnel  is  washed  repeatedly,  as  in  an  ordinary  filter. 

Diffusion  is  an  excellent  quantitative  method  for  the  investigation 
of  particle  size.  The  performance  of  diffusion  experiments  is,  how- 
ever, a  difficult  matter,  because  even  variations  in  temperature  may 
cause  material  errors. 

Dialysis.^ 

The  most  varied  apparatus  and  membranes  may  be  used  for 
dialysis.    An  apparatus  described  in  most  textbooks  and  used  in  many 

^  An  exhaustive  description  of  all  known  methods  of  dialysis  is  given  by  E. 
ZuNZ  in  Abderhalden's  Handb.  d.  biochem.  Arbeitsmethoden  3,  pages  165-189 
and  Supplement,  pages  478-485. 

89 


90  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

teaching  laboratories  is  the  one  originally  described  by  Graham.  A 
wide-mouthed  salt  bottle  A  (Fig.  9),  with  its  bottom  broken  ojff,  has 
a  pig  or  ox  bladder  or  a  piece  of  parchment  bound  about  the  neck; 
the  bottle  is  placed  with  the  membrane  downward,  in  a  vessel  of 

water  {By.     The  solution  to  be  dialyzed 

is  placed  in  the  bottle. 

Precautions:  Before  the  membrane  is  used, 
it  should  be  tested  to  see  that  it  can  be  made 
wet  by  water.  Greasy  animal  membranes 
must  be  rinsed  with  fresh  water  several  times, 
on  both  sides.  To  determine  whether  the 
Fig.  9.     A  simple  dialyzer.       membrane  leaks,   a  colored   solution    {e.g.,  a 

drop  of  colloidal  silver,  Utmus,  or  water  colored 
with  hemoglobin)  is  placed  in  A  and  allowed  to  remain  there  several  hours, 
without  putting  any  water  in  the  outer  vessel.  Colored  drops  will  pass  through 
at  points  of  leakage  (the  margin  where  the  membrane  is  bound  should  be  espe- 
cially watched) .  To  make  sure  that  the  drops  are  really  colored  solution  and 
not  pure  water,  they  should  be  absorbed  by  filter  paper. 

In  all  dialysis  experiments,  the  substance  under  examination  may  simulate 
colloidal  character  by  being  bound  or  adsorbed  by  the  dialyzing  membrane. 
Under  these  circumstances,  if  we  wish  to  determine  whether  the  solution  under 
examination  contains  colloidal  substances,  it  is  necessary  to  use  the  smallest 
possible  membrane  with  the  largest  available  amount  of  substance.  If  too  little 
substance  is  used,  it  may  all  become  bound  by  the  membrane,  and  in  spite  of  the 
fact  that  it  is  not  colloidal,  none  will  be  found  in  the  dialyzer.  But  if  so  much 
of  the  substance  under  investigation  is  used,  that  in  spite  of  any  possible  ad- 
sorption by  the  membrane,  plenty  still  remains  in  the  dialyzer,  then  an  examina- 
tion of  the  water  outside  wiU  settle  the  question. 

In  practice,  Graham's  dialyzing  apparatus  is  not  frequently  em- 
ployed, because  it  has  a  small  dialyzing  surface,  and  this  is  a  great 
disadvantage. 

In  order  to  bring  the  largest  possible  surface  into  contact  with 
the  surrounding  water,  there  are  used  either  whole  pig  bladders,  ox 
bladders,  fish  bladders,  or  the  commercial  parchment  thimbles.^ 

I  have  had  very  good  results  with  fish  bladders  (condoms)  which 
are  very  thin,  uniform  and  elastic,  though  unfortunately  they  are 
expensive.  Parchment  thimbles,  referred  to  above,  are  recommended 
for  the  dialysis  of  large  quantities  of  solution.  They  may  be  obtained 
in  all  sizes  and  in  all  lengths.  The  suspension  of  a  fish  bladder  is 
conveniently  accomplished  by  pressing  it  between  two  glass  rods 
which  are  held  together  by  rubber  bands  (cut  from  gas  tubing) ;  the 

1  With  organic  solvents,  instead  of  water,  alcohol,  benzol,  etc.,  must  of  course 
be  employed  and  the  membrane  (preferably  collodion)  is  previously  soaked  in 
these  fluids. 

^  These  are  on  sale  at  the  "Vereinigten  Fabriken  für  Laboratoriums-bedarf" 
Berlin,  Scharnhorst  Str.     (The  Kny  Scherer  Co.,  N.  Y.,  are  the  American  agents.) 


METHODS   OF   COLLOID   RESEARCH 


91 


Fig.  10.    A  fish  bladder 
(condom)  dialyzer. 


glass  rods  are  laid  over  a  tall  narrow  beaker,  in  which  the  water  is 
placed.  Parchment  tubes  are  suspended  in  a  similar  manner.  To 
avoid  tying  one  end  of  the  tube,  it  is  hung  up  in  U-fashion  so  that 
both  open  ends  are  pressed  together  by  the  glass  rods  (see  Fig.  10). 

Precautions:   In  filling  and  hanging  the  parchment  tubes  the  air  must  be  en- 
tirely pressed  out  before  fastening  the  ends,  for  should  water  dialyze  in,  consider- 
able pressure  will  develop,  which  may  burst  the  mem- 
brane.    All  the  precautions  given  on  page  90  must  be 
observed  (degreasing,  absorption,  etc.). 

Excellent  dialysis  membranes  may  be  pre- 
pared of  collodion  or  glacial  acetic  acid-collo- 
dion. Their  great  advantage  is  that  they 
may  be  prepared  in  any  desired  size,  shape 
and  degree  of  permeability,  and  are  easily 
sterilized. 

As  an  example,  we  shall  explain  how  to 
make  one  such  membrane.  A  test  tube  is 
dipped  into  collodion  and  then  allowed  to  drip, 
being  twirled  meanwhile,  and  when  it  has  skinned 
over,  the  whole  tube  is  quickly  immersed  in 
water.  In  a  short  time,  the  tube  is  circumcised 
at  a  height  which  will  give  a  membrane  of  the 
desired  length;  with  a  little  practice,  the  membrane  can  be  removed 
from  the  tube  with  ease.  The  membrane  may  be  formed  inside  the 
tube  also  by  rinsing  the  test  tube  with  collodion  or  glacial  acetic 
collodion  and  then  filling  it  with  water.  In  a  similar  manner,  spher- 
ical or  cylindrical  membranes  of  30  to  40  cm.  long  and  10  cm. 
diameter  may  be  made  (see  Fig.  11).  Such  sacs  may  be  fastened 
to  a  glass  tube  with  thread  or  collodion  so  as  to  form  a  water- 
tight joint.  The  membranes  are  best  preserved  in  water  to 
which  a  little  chloroform  has  been  added  to  prevent  the  growth  of 
moulds. 

In  making  his  collodion  sacs,  G.  Malfitano  ^  uses  glass  tubes  of  the 
shape  shown  in  the  accompanying  diagram,  which  are  rotated  by  a 
motor  to  secure  uniform  drying.  The  spherical  swelling  affords  an 
easier  removal  of  the  rim  (see  Fig.  12).  After  the  cut  is  made 
through  the  collodion  skin  at  the  equator  of  the  sphere  (— »),  it  is 
carefully  loosened  from  the  glass  and  turned  inside  out  or  the  rim  is 
fastened  to  a  large  glass  tube  which  is  exhausted,  thus  the  skin  is 
loosened  from  the  spherical  portion.  J.  Duclaux  uses  very  thin 
tubes,  about  1  cm.  in  diameter  and  1  meter  long,  so  as  to  get  the 
largest  possible  surface. 

^  According  to  a  personal  communication. 


92 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


G.  Malfitano  and  J.  Duclaux  use  their  sacs  chiefly  for  ultrafil- 
tration though  they  are  equally  useful  for  dialysis.  Biologists  fre- 
quently use  little  reed  sacks  for  dialyzing;  they  are  frail  though 
sterilizable,  but  their  capacity  is  small. 

Schleicher  and  Schüll  (Düren)  sell  dialyzing  thimbles.  They 
are  tubular,  closed  at  the  bottom,  moderately  firm,  made  of  parch- 


FiG.  11.    Collodion  sacs.     (A.  Schoef.) 


ment  and,  since  they  maintain  their  form,  they  are  useful  for  certain 
experiments.     They  are  employed  in  the  Abderhalden  test. 

R.  ZsiGMONDY  *=*  constructed  a  very  useful  "star  dialyzer."  ^  The 
hard  rubber  ring  B  (see  Fig.  13)  is  covered  by  a  membrane  (col- 
lodion, parchment  or  the  like)  and  is  placed  on  a  plate  A,  which 
carries  the  star-shaped  support.     Through  the  central  opening  in  the 

^  Obtainable  from  Robert  Mittelbach,  Göttingen. 


METHODS  OF  COLLOID  RESEARCH 


93 


plate,  water  flows  and  bathes  the  extensive  dialyzing  surface  of  the 
dialyzer  A. 

JoRDis  has  constructed  an  apparatus  resembhng  a  filter  press  for 
dialyzing  large  quantities,     A  number  of  wooden  rings  are  soaked  in 


/Ai 


^B^A 


Fig.  12.     Tube  for  the  preparation  of 
collodion  sacs.     (G.  Malfitano.) 


(R. 


13.     Star  dialyzer. 
Zsigmondy.) 

paraffin  and  then  both  surfaces  are  covered  with  parchment.  These 
rings  are  placed  in  an  apparatus  G  so  that  between  each  ring  with 
parchment  there  is  one  without  parchment.  They  are  made  tight 
with  rubber  washers.     The  individual   elements   of   the  filter  are 


iVooc/en  Ring,  I  cm.  thick       ^^ 


W=  Running  Water 
0=Diaiysing  Spaces 

Fig.  13a.     Continuous  dialyzing  apparatus  of  E.  Jordiss. 

Reproduced  from  Zeitschrift  fur  Electrochemie,  p.  677,  Vol.  VIII,  1902. 

pressed  together  with  wing  nuts  so  that  they  are  water-tight.  The 
solution  to  be  dialyzed  is  placed  in  the  rings  covered  with  parch- 
ment, the  water  in  the  intervening  rings  circulates  through  holes 
made  in  them  for  the  purpose,  see  Fig.  13a. 


94 


COLLOIDS  IN  BIOLOGY  AND   MEDICINE 


An  excellent  apparatus  ^  for  continuous  dialysis  which  also  permits 
a  concentration  of  the  dialyzate  has  been  constructed  by  Kopac- 
ZEWSKi.  He  apphes  the  idea  of  the  Soxhlet  extraction  apparatus. 
A  collodion  sac  prepared  as  described  on  page  91  is  placed  in  the  tube 
A  filled  with  water.  The  dialyzate  from  the  collodion  sac  may  be 
removed  either  through  the  cock  C  or  may  pass  either  at  once  or  drop 
by  drop  into  the  vessel  B.     If  B  is  heated,  the  water  vaporizes  and 


/^b 


a  S. 


KOPACZEWSKfS 
APPARATUS 


Fig.  13b. 


Fig.  14.    Dialyzing  filter.     (L.  Morochowetz.) 


passes  into  the  two  condensers  and  drops  again  through  a  and  6  into 
the  tube  containing  the  collodion  sac.  Since  with  biological  fluids 
the  temperature  must  remain  below  50°  C,  the  heating  is  done  under 
partial  vacuum  (reduced  pressure).  The  tube  is  emptied  through 
the  lateral  outlet  having  a  cock  c.  A  concentrated  dialyzate  is 
finally  obtained  in  the  vessel  B. 

Where  there  is  no  running  water  available  for  dialyzing,  the  dialy- 
zing filter  of  Leo  Morochowetz  may  be  employed.  Its  arrange- 
ment may  be  seen  in  Fig.  14.  The  funnels  may  be  obtained  from 
any  supply  house  and  the  parchment  filters  from  Schleicher  and 
ScHULL  (Düren). 

1  The  apparatus  may  be  obtained  from  Poulenc  freres,  122  Boulevard  St. 
Germain,  Paris,  France. 


METHODS  OF  COLLOID  RESEARCH 


95 


Dialysis  is  considerably  hastened  by  agitating  the  dialyzing  fluid. 
I  have  never  found  it  suggested  that  the  contents  of  the  dialyzer 
should  be  stirred  but  this  is  a  useful  procedure  with  non-foaming 
solutions.  F.  Hofme-ister  fastens  all  the  dialyzing  thimbles  to  a 
common  rod  which  he  rocks  up  and 
down  with  a  motor.  R.  Köhler  places 
fish  bladders  in  wide-mouthed  bottles 
which  he  closes  with  a  rubber  cork  and  a 
rubber  cap,  and  then  places  it  in  a  shak- 
ing machine;  to  prevent  twisting  off  the 
fish  bladder  at  its  neck,  he  inserts  sev- 
eral glass  rods  as  shown  in  Fig.  14a. 


Ultrafiltration. 


PUBBE/?  CAP 


RUBBER  STOPPER 


-FISH  BLADDER 


■GLASS  RODS 


Fig.  14a. 


H.  Bechhold  defines  ultrafiltration 
as  filtration  through  jelly  filters.  They 
serve  to  separate  colloid  solutions  from 
crystalloids  and  for  the  separation  of  colloid  mixtures  having  parti- 
cles of  different  size.  If  we  know  the  size  of  the  pores  in  an  ultra- 
filter,  ultrafiltration  affords  information  as  to  the  size  of  the  particles 
in  the  colloid  under  investigation. 

Ultrafilter.  For  ultrafiltration,  sac-like  membranes  may  be  em- 
ployed prepared  as  for  diffusion  experiments  (see  p.  91)  and  mounted 
as  shown  in  Fig.  11. 

A.  ScHOEP  *  increased  the  permeability  of  membranes  by  adding 
glycerin  and  castor  oil  to  the  collodion.  This  is  of  great  importance 
in  filtering  inorganic  colloids. 

This  sort  of  ultrafiltration  is  used  especially  in  France  (G.  Mal- 
FiTANO,  J.  DucLAUx),  but  it  is  of  limited  utility.  Filtration  occurs 
very  slowly  (a  few  cubic  centimeters  per  hour)  and  the  filters  can- 
not withstand  much  pressure,  so  that  their  usefulness  is  very 
limited. 

For  ultrafiltration,  H.  Bechhold**  used  pieces  of  filter  paper  im- 
pregnated with  jellies.  By  means  of  this  paper  support  the  filters 
acquire  great  strength  and  may  at  times  sustain  in  Bechhold's  ultra- 
filtration apparatus,  pressures  of  20  atmospheres  or  more.  Since 
H.  Bechhold  discovered  that  the  premeability  or  tightness  of  the 
ultrafilters  depended  on  the  concentration  of  the  jellies  used  in  pre- 
paring them,  it  is  possible  to  make  filters  with  pores  of  any  desired 
size.     The  filters  may  be  purchased  ready-made.^ 

1  Schleicher  and  SchüU,  in  Düren,  market  Bechhold's  ultrafilters  in  aluminium 
boxes  which  contain  10  filters  filled  with  water  and  sealed  with  a  rubber  ring, 
(diam.  9  cm.).     This  firm  keeps  in  stock  six  kinds,  of  different  porosity. 


96 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Since  some  may  desire  to  make  the  filters,  brief  direction  for  doing 
so  are  given. ^ 

The  most  useful  kinds  of  filter  paper  are  No.  566  and  No.  575  of 
Schleicher  and  Schüll,  These  are  cut  into  discs  of  9  cm.  in 
diameter  and  impregnated  with  the  jellies  under  atmospheric  pres- 
sure in  a  glass  trough  from  which  the  air  has  previously  been  ex- 
hausted, making  a  vacuum. ^ 

The  square  trough  T  (see  Fig.  15)  has  its  cover  ground  airtight. 
On  the  cross  bar  B  a  number  of  filter  papers  are  suspended.  The 
cover  C  has  two  openings;  through  No.  1  pass  two  tubes,  one  of 
which  leads  to  the  air  pump  A  and  the  other  to  the  pressure  gauge 


Fia.  15.    Trough  for  the  preparation  of  ultrafilters.     (H.  Bechhold.) 

m.  When  the  air  is  exhausted  from  the  trough,  the  fluid  jelly  is  al- 
lowed to  enter  through  the  funnel  F,  which  has  a  cock  and  a  tube 
leading  to  the  bottom,  until  sufficient  is  admitted  to  cover  the  filters. 
Then  the  valve  leading  from  the  funnel  is  closed  and  the  valve 
through  which  the  air  was  exhausted  is  opened  so  that  the  jelly  is 
forced  into  the  filters  under  atmospheric  pressure.  After  a  time 
(with  diluted  jellies  10  to  20  min.  with  concentrated  jellies  one  or 
two  hours)  the  cover  is  taken  off  and  the  rod  with  the  filters  is  re- 
moved from  the  fluids.  While  the  filters  are  draining,  they  are 
constantly  shaken.  Finally  the  v/hole  filter  is  rapidly  gelatinized  by 
plunging  it  into  a  suitable  fluid.     In  the  case  of  glacial  acetic  acid 

1  Given  in  detail  in  the  original  paper  of  H.  Bechhold.** 

2  May  be  obtained  from  the  Vereinigten  Fabriken  für  Laboratoriumsbedarf, 
Berlin. 


METHODS  OF  COLLOID  RESEARCH 


97 


collodion,  water  is  used.  If  gelatin  is  being  used,  the  entire  impreg- 
nation trough  must  be  placed  in  a  bath  of  lukewarm  water.  Gelatin 
filters  are  hardened  by  placing  the  filters,  still  moist  and  gelatinized 
in  the  air,  into  a  2  to  4  per  cent  ice  cold  formaldehyde  solution  and 
keeping  them  for  a  time  in  an  ice  box. 

The  filters,  however  prepared,  are  washed  several  days  in  running 
water  and  preserved  in  water  to  which  a  little  chloroform  has  been 
added  to  prevent  the  growth  of  mould. 

H.  Bechhold  generally  uses  glacial  acetic  acid  collodion,  a  solu- 
tion of  soluble  cotton  in  glacial  acetic  acid.^  By  diluting  with  glacial 
acetic  acid  the  solution  may  be  reduced  to  the  desired  concentration. 

If  non-aqueous  solutions  {e.g.,  benzol,  ether,  etc.)  are  to  be  ultra- 
filtered,  the  water  must  be  displaced  by  a  series  of  solvents.  (Water 
is  first  displaced  by  acetone,  the  latter  by  benzol,  and  so  on.) 


Fig.  16.     Ultrafilter.     (H.  Bechhold.) 


The  Ultrafiltration  Apparatus.  Very  porous  filters  are  permeable 
under  low  pressure  and  can  be  used  like  any  other  filter.  In  by  far  the 
largest  number  of  cases,  a  pressure  of  from  1  to  20  atmospheres  must 
be  exerted  to  obtain  any  filtrate  at  all.  For  this  purpose  H.  Bech- 
hold prepared  an  apparatus  which  in  its  simplest  form  is  shown  in 
Fig.  16  2;  Fig.  17  is  more  suitable  for  very  high  pressures.  Fig.  16 
consists  of  a  cylindrical  vessel  H  into  which  is  inserted  the  funnel  T. 
Between  the  lower  flanges  of  T  and  H,  the  disc  of  filter  paper  is 
pressed.     This  is  made  tight  by  the  two  rubber  rings  G,  G. 

To  protect  it  from  being  torn,  the  filter  lies  on  a  nickel  netting  or 
perforated  nickeled  plate  N  and  is  further  protected  from  bulging  un- 
der pressure  by  the  plate  P,  which  has  several  holes  in  it.  The  upper 
part  of  the  funnel  T  is  ground  conically  and  is  closed  by  means  of 

^  The  Chemischer  Fabrik  auf  Aktien  (vorm.  Schering),  Berlin,  prepares  to 
order  solutions  of  10  per  cent  collodion  and  2\  per  cent  potassium  carbonate, 
which  show  only  slight  tendency  to  contract  when  they  gelatinize. 

2  All  this  apparatus  is  manufactured  by  the  Vereinigten  Fabriken  für  Labora- 
toriumsbedarf, Berlin,  Scharnhorst  Str. 


98 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Fig.  17. 


Ultrafilter  for  high 
pressures. 


the  cover  D  with  a  conical  joint  and  a  rubber  washer.  By  turn- 
ing the  screw  cap  Sehr,  the  cover  above  as  well  as  the  filter  below 
is  tightened.  A  small  nipple  with  a  screw  thread  passes  through  the 
cover  and  to  this  the  pipe  from  the  pressure  chamber  is  attached. 

The  apparatus  shown  in  Fig.  17  is  chiefly 
used  for  pressures  above  10  atmospheres, 
and  is  closed  with  flanges.  Naturally 
this  is  more  bulky.  The  lettering  in 
Fig.  17  corresponds  with  that  of  Fig.  16, 
so  that  it  is  unnecessary  to  duplicate  the 
description.  An  apparatus  with  a  stirrer 
(also  on  the  market)  is  usually  prefer- 
able, because  filtration  is  relatively  more 
rapid  and  the  filtrate  is  more  uniform. 
The  omission  of  stirring  may  permit  a 
gel  layer  to  form  on  the  ultrafilter,  and 
this  gel  layer  may  then  act  as  a  filter 
itself.  It  is  especially  important  to  have 
the  packing  tight  against  high  pressures.  In  this  apparatus  the 
pressure  is  introduced  through  a  side  opening  because  the  stirrer 
occupies  the  central  one. 

The  Pressure.  The  pressure  may  be  produced  by  a  hand 
pump.  This  is  especially  useful  in  the  scientific  investigation  of  the 
action  of  filters,  where  fine  gradations  of  pressure  are  involved,  and 
where  prolonged  pressure  is  unnecessary.  In  practical  ultrafiltra- 
tion, it  is  preferable  to  use  a  steel  cylinder  containing  either  com- 
pressed air,  nitrogen  or  carbonic  acid,  etc.  Between  the  steel 
cylinder  of  the  ultrafiltration  apparatus,  a  reducing  pressure  valve  and 
two  manometers  are  introduced;  one  for  very  high  pressure  shows 
the  pressure  in  the  cylinder,  the  other,  beyond  the  valve,  the  lower 
pressure  in  the  ultrafiltration  apparatus.  A  second  reducing  pressure 
valve  near  this  manometer  permits  such  delicate  differences  in 
pressure  that,  in  my  opinion,  it  is  safe  to  use  this  arrangement  in- 
stead of  the  hand  air  pump  even  in  scientific  measurements. 

The  extensive  use  achieved  by  Bechhold's  ultrafiltration  has  led 
to  some  modifications  for  special  purposes.  Zsigmondy,  Wilke- 
DÖRFURT  and  Galecki  recommended  collodion  skins  for  analytical 
purposes.  They  placed  collodion  skins  on  a  Büchner  funnel  (Gooch 
filter,  i.e.,  porcelain  funnel  with  perforated  diaphragm)  and  thus 
filtered  off  coarse  colloids,  especially  inorganic  ones,  by  means  of  a 
tap-water  suction  pump.  This  arrangement  is  not  suitable  for 
pressures  above  one  atmosphere.  [J.  F.  McClendon  employs  alun- 
dun  thimbles  coated  with  collodion.  Tr.] 
The  essential  new  point  in  the  apparatus  devised  by  Burian  E. 


METHODS  OF  COLLOID  RESEARCH 


99 


Pribram  and  Kirschbaum  is  the  application  of  compressed  air  for 
stirring;  the  gas  which  supplies  the  pressure  enters  the  bottom  of  the 
vessel  through  a  perforated  spiral  and  thus  agitates  the  fluid.  I 
have  had  no  occasion  to  determine  whether  this  has  any  advantages 
over  the  mechanical  stirrer. 

The  Gauging  of  Ultrafilters.  It  is  important  in  many  cases  to 
have  a  measure  for  the  limits  of  effectiveness  of  ultrafilters,  as  in 
this  way  we  may  obtain  information  concerning  the  size  of  the  par- 
ticles of  the  colloid  under  investigation.  The  following  methods 
given  by  Bechhold  are  suitable  for  the  purpose: 

1.  Hemoglobin  Method:  A  1  per  cent  solution  of  hemoglobin 
(hemoglobin  scales,  Merck)  is  prepared  and  the  filter  is  tested  to 
see  if  it  permits  the  hemoglobin  to  pass  through.  If  the  hemoglobin 
is  retained,  the  filter  is  impermeable  to  most  inorganic  colloids 
(with  the  exception  of  freshly  prepared  silicic  acid).  The  degree  of 
permeability  of  the  filter  for  hemoglobin  may  be  recognized  by  the 
intensity  of  the  red  color  in  the  filtrate. 

H.  Bechhold  has  prepared  the  following  table  of  permeability 
for  ultrafilters,  which  is  arranged  in  the  order  of  the  diminishing 
size  of  the  particles  of  the  colloids  in  solution,  and  was  obtained  by 
using  ultrafilters  having  different  degrees  of  porosity. 


Suspensions. 

Prussian  blue. 

Platinum-sol,  Bredig. 

Ferric  oxid  hydrosol. 

Casein,  in  milk. 

Arsenic  sulphid  hydrosol. 

Gold      solution,      Zsigmondy, 

No.  4,  c.  40  fxjjL. 
Bismon,  colloidal  bismuth  oxid, 

Paal. 
Lysargin,  colloidal  silver,  Paal. 
Collargol,    silver,    v.  Heyden, 

20  /x/i. 
Gold,     solution,     Zsigmondy, 

No.  0,  c.  1-4  iJLfjL. 
1  per  cent  gelatin  solution. 


1  per  cent  hemoglobin  solution, 
molecular  weight  c.  16,000. 

Serum,  albumin,  molecular 
weight   5000   to    15,000. 

Diphtheria  toxin. 

Protalbumoses. 

Colloidal  silicic  acid. 

Lysalbinic  acid. 

Deutero  albumoses  A. 

Deutero  albumoses  B,  mol.  wt. 
c.  2400. 

Deutero  albumoses  C. 

Litmus. 

Dextrin,  mol.  wt.  c.  965. 

Crystalloids. 


2.  Air  Transpiration  Method.^  This  method  affords  approxi- 
mately absolute  values  for  the  largest  pores  of  an  ultrafilter.  It  is 
based  on  the  following  principle.     In  order  to  force  air  through  a 

1  Before  actually  undertaking  methods  2  and  3  the  original  paper  should  be 
consulted  (Bechhold*^),  as  the  details  cannot  be  abstracted. 


100  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

capillary  completely  immersed  in  and  wet  with  water,  a  certain 
pressure  is  necessary,  which  depends  upon  the  surface  tension  of 
water  against  air  (which  is  a  constant)  and  the  diameter  of  the 
capillary.  If  D  is  the  diameter  of  the  capillary,  p  the  pressure  in 
atmospheres  and  ß  the  capillarity  constant,  the  following  formula 
applies : 

D= ^ 

p.1.033  +  10' 

li  ß  =  7.7  at  18°,  we  obtain 

D  = 


30.8 


OP' 


p.1.033  +  10^ 

With  the  aid  of  this  formula,  the  smallest  diameter  of  the  pores 
in  question  may  be  calculated  from  the  least  pressure  necessary  to 
drive  air  through  the  pores  of  the  completely  wet  filter. 

The  practical  performance  of  the  experiment  is  as  follows:  The 
filtering  apparatus  is  turned  upside  down,  a  thin  layer  of  water  is 

placed  on  the  filter  (several  milli- 
meters high)  and  the  highest 
pressure  at  which  air  bubbles 
begin  to  escape  is  determined. 
Casing  \        "  The   diagrammatic   sketch,   Fig. 

18,  shows  the  filtering  apparatus 
in  normal  position  {T  =  funnel, 
F  =  ultrafilter,  A  =  air  intake).  Fig.  19  shows  it  in  the  position 
necessary  for  the  forcing  through  of  air.  Above  the  filter  there  is 
a  thin  layer  of  water. 

According  to  this  method  the  largest  pores  of  a  filter  which  just 
permits  hemoglobin  to  pass  through  have  a  diameter  50  to  99  ix/x. 

3.  Method  Based  upon  the  Rate  of  Transfusion  for  Water.  This 
method  affords  approximately  absolute  values  for  the  average  diam- 
eter of  the  pores  of  ultrafilters.  The  method  is  based  upon  the 
somewhat  indefinite  law  of  Poiseuille  for  the  passage  of  fluids  through 
capillary  tubes.^ 
D  =  diameter  of  the  pores. 

Q  =  amount  of  water  flowing  through  the  surface  F  under  the 
constant  pressure  S. 

R  is  the  ratio  between  the  empty  (water  containing)  space,  and 
that  filled  with  solid.  This  is  determined  from  the  percentage  of  dry 
material  in  the  jellies  (a  5  per  cent  filter  contains  full  and  empty 
space  in  the  proportion  of  5  to  95). 

1  Bechhold.*« 


METHODS  OF  COLLOID  RESEARCH  101 

L  is  the  length  of  the  capillaries  {i.e.,  not  smaller  than  the  thick- 
ness of  the  wet  filter). 

Ä;  is  a  constant  factor  dependent  on  temperature  and  kind  of  fluid. 
The  following  formula  applies : 

Q{R  +  I)L 


D  = 


k'S'F-R 


If  all  the  experiments  are  performed  under  the  same  conditions, 
the  formula  may  be  simplified,  because  ^ — ^ — ^  becomes  a  constant. 

For  the  practical  performance  of  this  experiment  two  persons  are 
required,  one  of  whom  regulates  the  pressure,  while  the  other  de- 
termines the  amount  of  water  filtered  over  a  certain  time,  fixed  by 
means  of  a  stop  watch. 

Under  the  apparatus  is  placed  a  funnel  which  has  a  rubber  tube 
with  a  pinch-cock  attached. 

The  ultrafiltration  apparatus  is  filled  with  water,  and  the  air  pres- 
sure is  raised  to  a  given  point.  The  pinch-cock  is  then  closed  so 
that  all  the  water  filtering  at  a  constant  pressure  is  caught  in  the 
funnel.  After  a  given  time  {e.g.,  one  minute)  has  elapsed,  the  entire 
pressure  is  instantly  released.  In  this  way  the  amount  of  water 
filtered  through  a  given  filter  in  a  given  time  is  measured.  If  we 
have  previously  performed  the  same  experiment  with  a  filter  paper 
which  has  pores  of  known  size,  one  which,  for  instance,  even  par- 
tially retains  blood  corpuscles  or  bacteria,  i.e.,  objects  which  are 
measurable  microscopically,  by  means  of  the  above  formula,  we 
can  estimate  the  average  size  of  the  pores  of  the  ultrafilter. 

With  this  method  ultrafilters  which  just  held  back  hemoglobin 
showed  the  average  diameter  of  their  pores  to  be  from  33  to  36  fx/j,. 

4.   Method  of  Emulsion  Filtration,  described  on  pp.  15  and  16. 

Adsorption  by  Filters.  In  ultrafiltration  experiments,  it  is 
necessary  to  avoid  errors  due  to  adsorption  on  the  part  of  the  filter. 
Accordingly,  as  a  preliminary  experiment,  it  is  advisable  to  shake  a 
portion  of  the  solution  with  a  shredded  filter.  If  the  content  of 
the  solution  is  practically  the  same  afrte  as  before  the  shak- 
ing, there  has  been  no  adsorption.  If  the  adsorption  introduces  an 
error  into  the  ultrafiltration  experiment,  it  is  necessary  to  use  a  dif- 
ferent jelly.  For  instance,  arachnolysin  is  very  strongly  adsorbed 
by  glacial  acetic  acid-collodion,  but  only  slightly  by  formol- 
gelatin. 

In  ultrafiltration  experiments  it  is  always  important  to  work 
quantitatively  and  to  test  what  remains  in  the  filter  as  well  as  the 
filtrate  obtained. 


102  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

[P,  A.  KoBER  has  devised  a  new  and  valuable  form  cf  ultiafilter 
based  on  the  principle  of  selective  dialysis  through  collodion  and 
evaporation  of  the  dialysate  (per-vaporation).  See  Jour.  Am.  Chem. 
Soc,  Vol.  XL,  No.  8,  po  1226,  et  seq.    Tr.] 

Applications  of  Ultrafiltration,  ultrafiltration  as  previously  men- 
tioned serves  to  separate  colloids  from  crystalloids.  It  can  fre- 
quently replace  dialysis,  having  the  advantage  of  rapidity  and 
permitting  separation  without  the  unavoidably  great  dilution  of  the 
dialysate. 

For  this  purpose  it  has  been  used  for  the  separation  of  globulin 
from  the  electrolytes  holding  it  in  solution,  and  the  products  of  the 
digestion  of  casein  by  pancreatin  (H.  Bechhold*^). 

The  most  important  recent  applications  of  ultrafiltration  are,  the 
separation  of  colloids  with  particles  of  different  sizes  (fractional  ul- 
trafiltration), and  the  determination  of  the  colloid  or  crystalloid 
nature  of  doubtful  substances.  We  refer  here  to  the  separation  of 
various  albumoses  by  H.  Bechhold,*^  the  researches  concerning 
the  nature  of  starch  solutions  by  E.  Fouard,*  and  those  concerning 
diastase  by  Pribram,  and  the  experiment  to  explain  fermentations 
in  the  absence  of  cells  by  A.  von  Lebedew,*  the  researches  of  Gros- 
ser on  milk  (see  pp.  174  and  350),  the  studies  of  Kirschbaum  on 
dysentery  toxin  which  are  still  unpublished,  as  well  as  those  of  H. 
Bechhold  on  the  separation  of  diphtheria  toxin  from  toxon.  By 
ultrafiltration,  Grosser  was  able  to  distinguish  boiled  from  unboiled 
milk  (see  p.  174). 

Ultrafiltration  is  of  especial  importance  in  the  study  of  equilibrium 
in  solutions,  because  in  this  method  there  is  no  change  in  the  balance 
of  crystalloid  and  colloid  portions  through  the  dilution  of  the  solu- 
tion. It  is  assumed  that  only  small  quantities  are  filtered,  that  the 
differential  is  in  some  way  ascertained  so  that  no  changes  in  concen- 
tration occur;  and  that  only  moderate  pressures  are  used  in  the 
case  of  solutions  containing  electrolytes  (see  p.  59).  The  numerous 
researches  on  iron  oxid  hydrosol  by  J.  Duclaux  and  G.  Malfitano 
depend  on  this,  as  does  the  work  of  R.  Burian*^  on  salt-albumin 
mixtures. 

Ultrafiltration  has  been  variously  employed  for  the  solution  of 
purely  biological  questions.  R.  Burian*^  has  employed  it  in  study- 
ing the  function  of  the  kidney  glomeruh,  and  H.  Bechhold*^  in  the 
question  of  "internal  antisepsis." 

Finally,  it  must  be  mentioned,  that  by  ultrafiltration  germ-free 
fluids  may  be  obtained,  as  well  as  optically  pure  water  suitable  for 
ultramicroscopic  experiments  (H.  Bechhold*^).  New  paths  have 
been  opened  to  the  study  of  filterable  infectious  agents  by  ultra- 
filtration (voN  Betegh). 


METHODS  OF  COLLOID  RESEARCH 


103 


Diffusion. 

Coefficients  of  diffusion  give  information  concerning  the  molecular 
weight  and  also  the  size  of  the  particles  of  a  substance  in  solution. 
Diffusion  in  aqueous  solution  is  the  simplest  method  for  such  investi- 
gations. The  length  of  time  necessary  for  such  experiments  intro- 
duces so  many  disturbing  factors  that,  where  possible,  diffusion  in 
a  jelly  is  to  be  preferred.  A  jelly  offers  a  means  of  separating  sub- 
stances having  different  rates  of  diffusion.  If  a  mixture  of  two  sub- 
stances remain  for  a  time  in  a  tube  partly  filled  with  a  jelly,  the  more 
difficultly  diffusible  substance  will,  for  the  most 
part,  remain  in  solution  and  can  be  poured  off, 
whereas  the  substance  easily  diffusible  will  to  a 
greater  extent  enter  the  deeper  layers  of  the  jelly. 
Diffusion  experiments  in  jellies  teach  us  the  prop- 
erties of  jellies  swollen  to  various  degrees,  both  in 
the  presence  of  crystalloids  and  in  their  absence. 

Diffusion  in  Aqueous  Solution.  The  greatest 
difficulty  lies  in  avoiding  agitation  not  only  when 
samples  are  being  taken  but  also  during  the  course 
of  the  experiments.  The  most  suitable  apparatus 
is  that  of  L.  W.  Öholm*  (see  Fig.  20),  with  which 
Herzog  made  his  experiments,  and  that  of 
Dabrowski.  The  latter  (see  Fig.  21a)  consists  of 
two  glass  vessels  A  and  B  (a  siphon  bottle  which 
has  been  divided  in  the  middle)  which  are  separated 
by  a  diaphragm  C.  This  diaphragm  is  a  glass  ring 
filled  with  glass  capillary  tubes  of  1  mm.  bore. 
The  interspaces  are  filled  with  celluloid. 

By  this  arrangement  currents  are  avoided  and 
a  very  considerable  diffusion  surface  is  obtained. 
The  solution  is  placed  in  A  and  diffuses  through  C  and  reaches 
B  from  which  samples  for  analysis  are  taken  from  time  to  time 
through  the  tube  F.  The  fluid  in  A  as  well  as  in  5  is  slowly  stirred 
by  the  stirrer  abd.  We  shall  return  to  the  consideration  of 
Dabrowski' s  experiments  on  the  diffusion  of  albumin  with  this 
apparatus  on  p.  72. 

In  the  extremely  slow  diffusion  of  colloids,  which  in  the  case  of 
the  experiments  of  R.  0.  Herzog  extended  over  more  than  two 
months,  absolute  sterility  is  essential.  Besides  having  sterile  vessels, 
the  solutions  are  also  sterilized  by  saturating  them  with  toluol  and 
layering  it  over  them.  The  addition  of  1/2  per  cent  sodium  fluorid 
solution  is  useful  also.     As  previously  mentioned,  the  vessels  must 


Hg 


Fig.  20.  Diffu- 
sion apparatus. 
(L.  W.  Öholm.) 


104  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

stand  in  a  perfectly  quiet  place;  instead  of  portable  water  baths, 
incubators  should  be  used,  or  if  these  are  not  placed  so  as  to  be 
free  from  vibration,  the  experiments  are  kept  by  preference  in  a 
cellar. 

These  diffusion  experiments  are  extremely  difficult,  but  may  yield 
absolutely  perfect  results,  as  has  been  shown  by  the  researches  of 
R.  O.  Herzog  and  H.  Kasarnowski.*  These  investigators  de- 
termined the  diffusion  coefficients  for  albumin  and  a  number  of 
enzymes  (see  p.  54  and  p.  190),  from  which  it  could  be  determined 
that  they  were  simple  substances.  On  the  other  hand  it  could  be 
shown  that  clupein  sulphate,  trypsin  and  pancreatin  were  mixtures 
of  different  substances.  Some  of  them  showed  various  diffusion 
layers  of  dissimilar  composition  (various  percentages  of  N,  in  clupein), 
and  with  other  mixtures,  products  of  different  origin  showed  different 
coefficients  of  diffusion  (trypsin,  pancreatin). 

Diffusion  in  a  Jelly.  A  jelly  acts  like  a  membrane.  It  has  the 
advantage  over  a  membrane  that  its  thickness  may  be  varied  at 
will,  but  the  disadvantage  that  it  is  usually  impossible  to  obtain  the 
diffused  substance  in  pure  form  so  that  the  diffused  substance  must 
be  examined  in  association  with  the  jelly.  Union  or  adsorption 
between  the  jelly  and  the  substance  under  examination  is  a  more 
disturbing  factor  than  in  the  case  of  a  membrane.  Diffusion  in  a 
jelly  has  the  great  advantage  over  the  diffusion  in  fluids,  above  de- 
scribed, that  it  is  not  disturbed  by  currents  or  the  almost  unavoidable 
shaking  during  the  experiment,  or  while  samples  are  being  taken. 

The  experiments  are  generally  performed  as  follows :  A  test  tube  is 
filled  one  third  to  one  half  full  with  a  very  dilute  jelly  (2  to  5  per  cent 
gelatin).  After  the  jelly  solidifies,  the  solution  to  be  investigated  is 
poured  upon  it,  and  the  test  tube  is  placed  in  an  ice  box.  After  a 
longer  or  shorter  time  (days,  weeks,  months)  some  of  the  substance 
will  have  diffused  into  the  jelly.  The  supernatant  fluid  is  now  poured 
from  the  jelly,  which  is  washed  with  a  suitable  fluid,  water,  physio- 
logical salt  solution  or  the  like.  The  jelly  is  now  examined  to  deter- 
mine the  course  of  diffusion,  taking  the  elapsed  time  into  consideration. 
Gelatin  and  agar  are  used  as  jellies.  In  many  cases,  inspection  shows 
the  extent  to  which  the  fluid  has  diffused  into  the  jelly  {e.g.,  with  dye- 
stuffs,  indicators  or  precipitation  reactions,  the  results  may  be  seen). 
For  example,  gelatin  which  has  been  mixed  with  red  blood  corpuscles 
may  have  tetanolysin  layered  above  it.  By  the  extent  of  the  hemo- 
lysis it  is  determined  how  far  the  tetanolysin  has  penetrated.  H. 
Bechhold*^  mixed  a  jelly  with  goat-rabbit  serum  and  layered  above 
it  a  solution  of  goat  serum.  The  appearance  of  a  white  ring  showed 
the  distance  that  the  precipitin  had  penetrated. 


METHODS  OF  COLLOID  RESEARCH 


105 


Precautions:  For  these  experiments  there  should  be  utiUzed  only  absolutely  pure 
gelatin  or  agar  which  has  been  dialyzed  at  least  two  or,  preferably,  four  or  five  days 
in  cold  running  water.  Commercial  gelatin  always  contains,  besides  certain  other 
impurities,  sulphurous  acid  which  is  used  as  a  bleach  in  its  manufacture,  and  as  a 
result  the  gelatin  reacts  acid  to  Utmus.  It  may  be  completely  freed  from  the 
acid  by  neutraUzation  with  NaOH  followed  by  sufficiently  prolonged  dialyzation 
in  running  water.  The  dried  gelatin  is  weighed,  wrapped  in  Unen  or  mull  and 
placed  in  a  trough  of  running  water.  After  purification  the  swollen  gelatin  is 
very  carefully  removed  from  the  cloth  and  weighed  to  see  how  much  water  it  has 
taken  up.  By  the  addition  of,  or  the  evaporation  of  water,  the  jelly  is  brought 
to  the  desired  concentration  and  filtered  through  a  jacketed  filter.  This  is 
usually  a  sufficiently  accurate  method.  For  absolutely  exact  investigation,  a 
measured  quantity  of  the  moist  gelatin  must  be  weighed  before  and  after  it  has 
been  dried  at  105°  C.  If  working  with  other  than  pure  aqueous  solutions,  such 
as  with  substances  which  require  physiological  salt  solution  to  dissolve  them, 
the  gelatin  or  agar  must  contain  the  required  amount  of  salt,  if,  for  instance,  the 
diffusion  of  globuUn  solution  is  desired.  Since  diffusion  experiments  with  col- 
loids always  extend  over  a  considerable  time,  the  test  tubes  must  be  closed  with 
paraffined  corks  or  rubber  stoppers. 

In  such  cases  quantitative  measurements  may  be  made  with  a 
ruler,  by  placing  the  zero  at  the  meniscus  of  the  jelly  or  by  means 
of  cathetometer.     Tubes  with   an   engraved   scale  as  arranged  by 


yju^ 


J 


J 


Fig.  21.    Diffusion  tube. 


Fig.  21a.     Dabrowski's 
diffusion  apparatus. 


Stoffel-Pringsheim*  (see  Fig.  21)  are  convenient.  The  graduated 
tube  is  filled  with  jelly  and  the  solution  is  poured  into  the  extensions 
which  are  ground  on  water  tight.  For  accurate  measurements  the 
same  rules  are  used  as  in  similar  physical  measurements. 

If  the  diffusion  into  the  jelly  is  not  associated  with  a  visible  change, 
tiie  jelly  is  removed  from  the  glass  by  placing  it  in  hot  water  until 
the  periphery  melts,  so  that  the  cy finder  of  jelly  may  be  gently 
pu-shed  out  of  the  tube.  The  jeUy  cyfinder  is  then  cut  into  layers 
of  measured  thickness  which  are  studied  by  chemical,  biological  or 
animal  experiments  as  to  their  content  of  diffused  substance.  In 
this  way  Sv.  Arrhenius*  and  Th.  Madsen  have  determined  the 
diffusion  constants  of  diphtheria  toxin  and  antitoxin  and  of  tetanoly- 
sin  and  antitetanolysin. 


106 


COLLOIDS  IN  BIOLOGY  AND   MEDICINE 


In  order  to  remove  the  gelatin  cylinder  easily,  H.  Bechhold  and 
J.  ZiEGLER  coated  the  interior  of  the  test  tube  with  a  lining  capsule 
of  parchment,  paraffined  paper,  pergamyn  or  the  like,  so  that  the 
paper  is  closed  below;  on  the  side,  it  is  closely  adherent  to  the  glass, 
while  above  it  projects  about  1  cm.  above  the  rim.  This  paper  lining 
is  filled  with  gelatin,  allowed  to  cool  quickly  and  removed  with  the 
gelatin  at  the  end  of  the  experiment.  The 
gelatin  cylinder  is  sliced  after  unwrapping  the 
paper. 

One  might  imagine  that  instead  of  determin- 
ing the  quantity  of  substance  which  had  diffused 
into  the  jelly,  i.e.,  the  diffusion  path,  the  percen- 
tage of  substance  that  has  been  lost  by  the  re- 
maining fluid  could  be  determined.  For  inves- 
tigations of  colloids  this  method  is  not  to  be 
recommended,  because  with  the  slight  diffus- 
ibility  of  colloids,  the  loss  of  substance  and  the 
limits  of  error  approach  each  other  closely. 

The  experiments  of  Voigtländer,  placing 
scales  of  glue  in  solutions  and  determining  the 
amount  of  the  dissolved  substance  that  they 
took  up,  are  not  suitable  for  use  with  colloid 
material. 

R.  Liesegang**  has  developed  a  special 
method.  He  covers  a  plate  with  a  jelly  and 
puts  on  it  drops  of  a  solution  which  diffuse  in 
rings.  The  method  is  especially  suitable  for 
qualitative  studies.  If  the  jelly  is  impregnated 
with  a  substance  which  forms  precipitates 
with  the  diffusing  solution,  structures  appear 
whose  form  and  growth  may  be  beautifully 
Fig.  22.  Osmometer  of  studied.  Instead  of  aqueous  solution,  sheets  of 
Biltz  and  Von  Vegesack.  ^^^y  ^^y  ^g  placed  on  the  gelatin  layer. 

Diffusion  and  capillary  ascension  in  filter  paper  (which  must  be  ab- 
solutely clean)  may,  under  certain  circumstances,  give  useful  quali- 
tative information. 

Osmotic  Pressure. 

While  in  the  case  of  crystalloids,  indirect  methods  of  determining 
the  osmotic  pressure  are  used  (lowering  the  freezing  point  or  raising 
the  boiling  point),  in  the  case  of  substances  lying  at  the  border 
line  of  colloids,  the  direct  osmotic  method  is  most  useful.  W. 
Biltz  and  A.  von  Vegesack*  constructed  an  osmometer  whose  main 


METHODS  OF  COLLOID  RESEARCH  107 

feature  consists  of  a  collodion  membrane  (see  Fig.  22)  protected  by 
a  platinum  wire  netting.  In  the  case  of  dextrins  which  lie  on  the 
border  line  between  colloids  and  crystalloids,  W.  Biltz  decreased  the 
permeability  of  the  membrane  by  adding  cupric  ferrocyanid.  He 
filled  the  collodion  sac  with  1  per  cent  potassium  ferrocyanid  solution 
and  placed  it  in  1  per  cent  copper  sulphate  solution.  After  twenty- 
four  hours  he  washed  the  sac  for  twenty-four  hours  in  running  water. 
The  method  of  procedure  recommended  by  Fouard,  impregnation 
with  tannin  and  gelatin  and  subsequent  tanning  with  sublimate,  has 
not  proven  effective,  according  to  Biltz.  Above  the  collodion  mem- 
brane is  a  glass  cap  with  a  vertical  tube.  The  union  of  netting  and 
cap  is  at  6.  The  fluid  is  mixed  by  an  electromagnetic  stirrer  c.  The 
electrodes  d  permit  the  measurement  of  the  conductivity.  The  entire 
instrument  is  placed  in  a  thermostat.  Readings  of  the  rise  in  the 
tube  are  made  with  a  cathetometer. 


Osmotic  Compensation  Method 

This  method  determines  whether  crj^stalloids  present  in  a  colloidal 
solution  are  free  or  in  any  way  bound,  e.g.,  adsorbed.  L.  Michaelis 
and  P.  RoNA*2  have  developed  this  method  in  their  attempt  to  solve 
the  question  whether  the  grape  sugar,  always  present  in  the  blood 
and  which,  strange  to  say,  does  not  pass  through  the  kidney,  is  free 
or  bound  in  any  way.  For  this  purpose  we  place  the  fluid  to  be  in- 
vestigated (in  this  instance  blood)  in  a  fish  bladder  and  suspend  it 
in  a  glass  cylinder  containing  an  isotonic  fluid  (in  this  case  water 
with  0.95  per  cent  NaCl).  To  the  surrounding  fluid,  in  a  series  of 
experiments,  there  is  added  varying  quantities  of  the  crystalloids  in 
question.  In  this  instance  sugar  is  added ;  we  shall  continue  to  describe 
the  above  experiment  as  an  example.  If  more  sugar  has  been  added 
than  is  present  in  the  blood  it  will  diffuse  into  the  blood;  the  sugar 
content  of  the  surrounding  fluid  will  decrease.  If  very  little  or  no 
sugar  is  added,  the  sugar  will  diffuse  from  the  blood  into  the  sur- 
rounding fluid.  This  will  occur  whether  the  sugars  in  the  blood  are 
free,  osmotically  active  or  even  if  a  portion  is  adsorbed.  In  the 
latter  case  the  free  sugar  will  at  first  diffuse  away  so  that  the  balance 
between  the  adsorbed  and  the  free  sugar  is  disturbed;  previously 
adsorbed  sugar  may  become  free  and  likewise  diffuse  away.  The 
sugar  content  of  the  outer  fluid  remains  constant,  only  if  it  accu- 
rately expresses  the  free  sugar  content  of  the  blood.  If  the  total 
sugar  has  been  determined  previously,  we  may  calculate  what  per- 
centage is  adsorbed  or  otherwise  bound,  and  what  proportion  is 
free  or  osmotically  active.     With  this  method,  L.  Michaelis  and 


108  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

P.  RoNA  determined  that  the  entire  sugar  in  the  blood  serum  is  in 
free  solution.  In  analogous  ways  these  authors  investigated  the 
conditions  of  union  between  calcium  and  the  casein  of  milk. 


Surface  Tension. 

Because  of  its  sensitiveness,  the  measurement  of  surface  tension 
is  of  the  greatest  significance  for  colloid  investigation.  As  far  as  I 
know  at  present,  there  is  no  case  in  which  the  measurement  of  these 
factors  has  led  to  the  solution  of  any  problem.  This  is  due  to 
the  fact  that  even  traces  of  other  substances,  especially  colloidal  sub- 
stances, markedly  influence  the  surface  tension  because  they  are  forced 
to  the  interfaces.  According  to  J.  Traube,  HgCl2  in  a  dilution  of 
one  in  three  million  may  be  detected  in  dye  solutions.  For  this  reason, 
measurements  of  surface  tension  are  excessively  sensitive  and  are 
subject  to  certain  errors. 

There  are  two  essentially  different  groups  of  methods:  (a)  static, 
(h)  dynamic } 

(a)  Static  Methods  (the  rise  of  a  fluid  in  a  capillary;  the  def- 
ormation of  an  air  bubble  in  a  fluid)  show  the  condition  of  the  de- 
veloped surface,  (b)  Dynamic  methods  (the  weight  or  number  of 
falling  drops;  the  pressure  necessary  to  force  air  through  a  capillary 
dipped  in  a  fluid)  show  the  condition  of  a  nascent  surface. 

These  two  methods,  especially  with  colloids,  give  fundamentally 
different  values  because  the  interior  of  a  fluid  has  a  very  different 
composition  from  the  surface,  and  a  considerable  time  always  elapses 
before  the  surface  has  assumed  its  normal  properties. 

As  yet,  because  they  are  the  simplest  to  perform,  only  the  capillary 
ascent  and  the  falling  drop  methods  have  been  used  for  biological 
studies. 

The  measurement  of  the  height  ascended  in  filter  paper  which  has 
been  used  especially  by  Goppelsröder  in  his  numerous  investigations 
on  alkaloids,  dyestuffs  and  other  organic  substances  ^  may  be  counted 
a  dynamic,  rather  than  a  static  method,  since  in  this  porous  material 
with  the  ascent  and  evaporation,  new  surfaces  are  continually  formed. 
Filter  paper  offers  a  very  useful  method  for  demonstration  purposes. 
Thus,  it  shows  why  alcoholic  solutions  and  soap  tinctures  rather  than 
aqueous  solutions  are  adapted  to  disinfection  of  the  skin  (according 
to  Bechhold)  (see  p.  404). 

1  Detailed  descriptions  of  the  methods  are  to  be  found  in  Ostwald-Luther's 
Physico-Chemischen  Messungen  (Leipzig,  1910),  and  in  G.  Quincke,  Poggen- 
dorff's  Annalen  d.  Physik,  139,  1-89  (1870). 

2  KoUoid  Zeit.,  Vol.  4,  pp.  41,  94,  191. 


METHODS  OF  COLLOID  RESEARCH  109 

J.  Traube  has  used  the  falUng  drop  method  with  his  stalagmom- 
eter  for  numerous  researches.  M.  Ascoli  has  hkewise  used  it  in  his 
meiostagmin  reaction  in  cancer.  [Clowes  employed  it  in  his  studies. 
See  p.  39.     Tr.] 

A  given  quantity  of  fluid  volume  is  sucked  up  into  the  stalagmom- 
eter  tube  and  the  number  of  drops  required  to  empty  it,  dropping  it 
drop  by  drop,  are  counted. 

The  stalagmometer  is  an  instrument  which  requires  most  careful  manipulation 
to  obtain  reUable  results.  It  is  especially  important  to  keep  it  scrupulously 
clean,  rinsing  frequently  with  distilled  water  followed  by  hot  potassium  hydrate 
solution.  The  apparatus  is  then  placed  over  night  in  a  hot  mixture  of  concen- 
trated sulphuric  acid  and  potassium  bichromate.  Before  use  it  is  again  thor- 
oughly rinsed  with  distilled  water.  The  dropping  surface  must  be  absolutely 
horizontal;  this  is  accomplished  by  placing  it  on  a  stand  which  can  be  adjusted 
in  all  directions.  There  must  be  no  bubbles  on  the  dropping  surface  or  in  the 
tubes.  Before  each  initial  measurement  the  fluid  to  be  measured  must  be  sucked 
up  and  allowed  to  flow  out  again.  The  number  of  drops  of  the  fluid  is  com- 
pared with  the  number  of  the  same  volume  of  water.  The  speed  of  flow  is  so 
gauged  that  no  more  than  20  drops  fall  in  a  minute.  This  is  best  regulated  by  a 
screw  clamp  on  a  rubber  tube  slipped  over  the  upper  end  of  the  instrument. 

J.  L.  R.  Morgan  has  devised  a  splendid  apparatus  for  determining 
the  weight  of  falling  drops  and  with  it  he  has  measured  the  surface 
tension  of  many  substances. 

There  are,  however,  valuable  contributions  to  the  utilization  of 
surface  tension  (milk  investigations  by  H.  Zangger;  meiostagmin 
reaction  of  M.  Ascoli). 

The  separation  of  colloids  and  crystalloids  hy  foaming  as  well  as  the 
separation  of  colloids  of  different  surface  tension  is  described  on 
page  35. 

Adsorption. 

Adsorption  experiments  may  be  employed  for  various  purposes. 
They  may  be  used  to  determine  the  distribution  of  a  dissolved  col- 
loid between  solvent  and  adsorbent,  thus  constituting  the  determi- 
nation of  a  physical  constant.  In  such  a  case  an  absolutely  chemically 
indifferent  substance,  e.g.,  charcoal,  is  chosen  as  absorbent.  Ad- 
sorption presents  a  suitable  means  of  determining  the  nature  of  the 
electric  charge  of  a  dissolved  colloid.  Positively  charged  colloids 
are  adsorbed  particularly  strongly  by  electronegative  suspensions 
(e.g.,  kaolin,  mastic  suspensions);  negatively  charged  colloids  are 
strongly  adsorbed  by  positive  suspensions  (e.g.,  iron  oxid,  clay). 
Occasionally  it  is  of  interest  to  determine  the  properties  of  a  gel 
when  it  is  used  as  an  adsorbent. 

If  in  all  cases  there  occurred  pure  adsorption,  whereby  a  dissolved 


110  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

substance  is  taken  up  by  a  solid  one  with  which  it  is  shaken,  the 
accurate  determination  of  adsorption  constants  would  be  of  the 
greatest  value.  They  would  then  be  natural  constants  of  the  same 
class  as  the  boiling  points,  melting  points,  etc.,  which  definitely  deter- 
mine the  nature  of  the  substances  under  examination.  Unfortunately 
this  is  not  the  case.  Chemical  phenomena  and  unexplained  factors 
complicate  the  pure  adsorption  phenomena,  so  that  at  present,  in 
biological  questions,  it  is  only  of  value  to  determine  whether  ad- 
sorption is  the  predominating  force.  Investigations  in  this  field 
are  of  great  importance.  Before  the  advent  of  physical  chemistry 
and  even  now,  in  biological  chemistry,  it  was  usual  to  search  for 
"  pure  "  substances,  and  to  illustrate  a  phenomenon  by  a  chemical 
equation.  Adsorption  experiments  have  frequently  made  it  clear 
to  us  that  in  a  given  case  such  chemical  equations  do  not  and 
could  not  exist.  The  studies  of  H.  Wislicenus  on  lignin  (see  p. 
248),  and  on  the  dyeing  process  by  W.  Biltz  and  H.  Freundlich, 
are  selected  from  among  many  other  classical  examples. 

Adsorption  experiments  for  determining  distribution  are  performed 
by  shaking  equal  quantities  by  weight,  of  the  most  indifferent  solid 
substance  obtainable  or  a  gel  (charcoal,  cellulose)  with  various  dilu- 
tions of  the  dissolved  substance.  The  amount  of  the  substance  ad- 
sorbed is  usually  ascertained  from  the  solution.  It  is  first  determined 
how  much  active  substance  is  contained  in  a  unit  volume  of  the 
solution,  which  is  then  examined  to  see  how  much  has  been  removed 
by  the  adsorbent;  the  difference  gives  the  quantity  adsorbed.  Thus 
H.  Wislicenus  determined  the  total  solids  in  the  cambial  sap  of  the 
beech,  before  and  after  shaking  it  with  cellulose,  and  found  by  taking 
the  difference  in  weight  the  amount  of  colloid  that  was  adsorbed. 

In  individual  instances  the  quantity  adsorbed  was  determined 
from  the  adsorbent.  B.  W.  Roux  and  Yersin  treated  diphtheria 
toxin  with  freshly  precipitated  calcium  phosphate;  they  then  washed 
the  calcium  phosphate  well  and  injected  it  into  guinea  pigs.  A  de- 
termination by  means  of  the  adsorbent  instead  of  the  fluid  I  con- 
sider erroneous  in  principle,  because  it  has  been  shown  repeatedly, 
in  well-controlled  experiments,  that  the  adsorbed  substance  under- 
goes changes  at  the  surface  of  the  adsorbent. 

The  fact  that  a  portion  of  the  dissolved  substance  is  removed  from 
the  fluid  by  a  solid  substance  with  a  large  surface  does  not  prove 
that  adsorption  has  taken  place.  If,  for  example,  1  gm.  cellulose 
always  removes  from  a  solution  the  absolutely  identical  quantity  of 
the  dissolved  substance,  irrespective  of  the  concentration  of  the 
solution,  we  would  in  all  probability  be  dealing  with  a  chemical  phe- 
nomenon.    If  the  proportion  between  the  adsorbed  substance  and 


METHODS  OF  COLLOID  RESEARCH  111 

i 
the  amount  still  in  solution  remains  constant  over  various  dilutions, 

we  may  assume  that  the  cellulose  forms  a  solid  solution  with  the  sub- 
stance in  question.  Adsorption  probably  exists  only  if  the  cellulose 
takes  up  almost  everything  from  a  very  dilute  solution  and  if  the 
absorbing  power  of  the  cellulose  is  markedly  decreased  with  increased 
concentration  of  the  solution;  this  condition  is  frequently  observed 
with  dye  solutions.  Thus  we  may  make  shaking  experiments  with 
solutions  of  the  concentration,  0.1,  0.2,  0.3,  etc.,  in  which  0.1  denotes 
any  arbitrary  standard. 

It  must  be  determined  first  whether  an  equilibrium  exists  at  all. 
For  this  purpose  a  given  quantity  of  adsorbent  is  shaken  with  the 
solution,  for  example,  with  100  c.c.  In  a  second  experiment  an 
equal  quantity  of  adsorbent  is  shaken  with  half  the  quantity,  50  c.c. 
of  a  solution  twice  the  strength.  It  is  then  diluted  to  100  c.c.  and 
shaken  again  until  an  equilibrium  is  reached.  If  there  is  an  equilib- 
rium, the  final  concentration  of  the  solution  in  the  first  case  is  the 
same  as  in  the  second.  If  there  are  material  differences,  the  process 
may  nevertheless  be  considered  an  adsorption,  but  it  is  complicated 
by  other  phenomena  as  explained  on  page  27  et  seq. 

If  it  is  unnecessary  to  determine  constants,  the  simplest  proce- 
dure is  to  chart  the  values  found  on  a  rectangular  system  of  co- 
ordinates (millimeter  paper).  As  ordinate  is  taken  the  amount  of 
the  material  that  is  being  investigated  which  is  taken  up  by  1  gm. 
of  adsorbent  (cellulose,  charcoal  or  the  like) ;  as  the  abscissa,  the 
amount  which  remains  in  solution  after  the  adsorption;  so  that  the 
curve  shows  the  ratio  between  the  amount  of  substance  in  the  solu- 
tion and  the  amount  that  is  adsorbed.  It  is  easy  to  determine  from 
the  characteristic  form  of  the  curve  whether  an  adsorption  has  oc- 
curred.    (See  p.  22.) 

The  determination  of  adsorption  curves  and  constants  is  explained 
in  detail  on  page  22  et  seq. 

It  is  of  greatest  importance  that  the  adsorbent  be  absolutely  pure.  Many  in- 
vestigators have  failed  in  this  and  many  contradictory  results  may  be  attributed 
to  it.  The  adsorbents  are  treated  with  acids,  alkaUes,  alcohol,  ether  and  benzol 
according  as  their  nature  permits  (charcoal,  diatomaceous  earth  or  kieselguhr, 
fibrin,  etc.)-  In  view  of  the  fact  that  these  substances  are  themselves  more  or 
less  adsorbed,  it  is  necessary  to  remove  them  by  prolonged  constant  treatment 
with  large  quantities  of  the  dispersing  substance,  usually  water. 

Although  temperature  and  time  do  not  play  as  important  a  role  as  in  other 
physico-chemical  processes  it  is  important  to  keep  temperature  and  time  con- 
stant. In  most  cases  the  adsorption  balance  is  reached  in  about  one-half  hour 
so  that  it  is  always  fairly  safe  to  allow  an  hour. 

It  is  usual  to  shake  the  adsorbent  with  the  solution,  but  it  must  not  be  over- 
looked that  there  are  substances  which  are  changed  by  the  mere  shaking  (see 
Inactivation  by  Shaking,  p.  34). 


112 


COLLOIDS  IN  BIOLOGY  AND   MEDICINE 


A  second  disadvantage  of  the  shaking  is  that  the  adsorbent  is  thereby  still 
further  broken  up  and  its  surface  thus  permanently  increased.  When  large 
quantities  of  colloid  are  in  solution,  there  is  a  counterbalancing  error  in  that  the 
adsorbent  becomes  coated  with  a  layer  of  coUoid  which  thus  diminishes  the  ac- 
tive surface.  Though  these  errors  are  small  in  the  case  of  adsorbed  crystalloids, 
in  the  case  of  true  colloids  they  become  quite  considerable.  To  eliminate  these 
two  disadvantages,  H.  Wislicentjs  and  W.  Muth*  have  developed  a  method 
which  they  caU  the  siphon  (or  filter)  process.  In  this  method  a  solution  of  con- 
stant strength  comes  repeatedly  in  contact 
with  the  adsorbent.  The  process  is  as  fol- 
lows: a  tube  is  filled  with  washed  clay  or 
other  adsorbent  and  in  connection  with  a 
separatory  funnel,  forms  a  siphon.  The 
solution  to  be  studied  is  poured  into  the 
funnel  and  very  slowly  filters  through  the 
adsorbent.  The  apparatus  (see  Fig.  23)  is 
entirely  practical.  In  the  strict  scientific 
sense,  however,  equilibria  are  not  obtained 
with  it. 


Fig.  23.     Apparatus  for  adsorption 
analysis.     (H.  Wislicenus.) 


Before  determining  the  content  of 
the  solution  after  adsorption,  the  ad- 
sorbent must  he  removed.  Filtration  is 
rarely  suitable  because  the  filter-paper 
itself  adsorbs.  In  any  event  the  filter 
used  should  be  very  small,  and  the 
quantity  of  fluid  to  be  filtered  as  large 
as  possible.  Centrifugation  is  the 
most  practical  method.  The  fluid 
may  be  poured  or  pipetted  from  the 
adsorbent  which  has  been  deposited. 
The  determination  of  the  content  of  the  solution  before  and  after 
adsorption  varies  so  much  in  accordance  with  the  nature  of  the  sub- 
stance under  investigation,  that  it  is  hardly  possible  to  formulate 
general  rules.  The  simplest  procedure,  when  it  is  possible,  is  to  de- 
termine the  weight  of  a  given  volume  after  evaporation,  or  the  solu- 
tions may  be  titrated.  In  other  cases  suitable  physical  or  biological 
methods  must  be  employed  (animal  experiment,  hemolysis,  agglu- 
tination, etc.). 

To  determine  the  electric  charge  of  a  colloid  by  adsorption,  we 
choose  for  adsorbent,  a  suspension  of  a  substance  having  the  most 
pronounced  electrical  charge.  Electropositive  iron  oxid  or  alumina 
gel  removes  electronegative  colloids  from  solution.  Electronega- 
tive diatomaceous  earth  (kieselguhr),  kaolin  or  mastic  suspensions 
(obtained  by  dropping  an  alcoholic  solution  of  mastic  into  water) 
attract  electropositive  colloids.      As  has  been  said,  the  charge  of 


METHODS  OF  COLLOID  RESEARCH  113 

natural  colloids  depends  chiefly  upon  their  reaction.  Experiments 
are  therefore  performed  with  very  faintly  acid,  very  faintly  alkaline 
and  neutral  reactions.  Because  many  substances  are  destroyed  in 
alkaline  or  acid  solutions,  it  is  necessary  to  make  appropriate  pre- 
liminary tests.  Measurements  of  the  amount  contained  before  and 
after  adsorption  determine  the  character  of  the  particular  colloid. 
In  this  way  L.  Michaelis  investigated  a  number  of  ferments  (see 
p.  186). 

In  the  border  land  between  adsorption  and  chemical  combination 
belong  the  studies  of  staining,  which  open  to  the  histologist  a  wide 
field  for  the  application  of  colloid-chemical  knowledge. 

Internal  Friction. 

As  has  been  shown,  especially  by  the  investigations  of  Wolfgang 
Pauli  on  albumin,  the  internal  friction  or  viscosity  serves  to  give 
valuable  information  concerning  changes  in  the  condition  in  colloidal 
solutions. 

In  the  case  of  hydrophile  colloids  an  increase  of  viscosity  usually 
indicates  an  hydration. 

The  relative  internal  friction  is  usually  determined  by  taking  that 
of  water  at  the  same  temperature  as  equal  to  1.  This  is  usually 
done  by  allowing  a  given  amount  of  fluid  to  flow  from  a  capillary 
tube,  taking  the  time  with  a  stop  watch.  If  the  rate  of  flow  for  water 
has  been  previously  determined,  the  relation  between  the  two  gives 
the  relative  internal  friction. 

Wilhelm  Ostwald  constructed  a  well-adapted  apparatus  (described 
in  Ostwald-Luther's  "Textbook  and  Manual  for  the  Performance 
of  Physico-chemical  Measurements,"  which  see  for  details).  The 
colloid  investigator  should  not  work  with  capillaries  that  are  too  fine, 
because  his  fluids  are  usually  very  viscous.  The  maintenance  of  a 
constant  temperature  is  of  especial  importance,  and  therefore  it  is 
necessary  to  employ  a  transparent  thermostat.  Furthermore  the 
specific  gravity  must  also  be  taken.  Ostwald-Sprengel's  Pyknom- 
eter may  be  used. 

In  biological  investigations  it  is  occasionally  necessary  to  work 
with  very  small  amounts  of  fiuid.  Special  apparatus  has  therefore 
been  devised  so  that  but  one  or  two  drops  may  suffice  for  a  viscosity 
determination.  The  apparatus  of  Hirsch  and  Beck,  thoroughly 
described  by  P.  T  Koranyi  and  A.  v.  Richter,  "  Physical  Chemistry 
and  Medicine  II,"  p.  27  et  seq.,  is  frequently  used.  The  apparatus 
of  H.  A.  Determan  is  very  simple;  as  seen  from  Fig.  24,  it  resembles 
an  hour  glass.     The  capillary  has  at  either  end  an  enlargement,  and 


114 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


■Thermome^-er 


ffffm 


^; 


then  a  constriction  as  well  as  markings.  This  tube  is  placed  in  a 
large  glass  shell  which  can  revolve  on  its  axis  and  has  a  thermometer 
inserted.  Since  the  apparatus  may  be  turned  upside  down  like  an 
hour  glass,  it  is  possible  to  take  several  successive  readings  from  the 
same  quantity  of  fluid.  Determan  employs  it  chiefly  for  the  de- 
termination of  viscosity  in  uncoagulated  blood.     For  this  purpose 

he  places  a  trace  of  hirudin  on 
the  unbroken  skin,  preferably  on 
the  lobe  of  the  ear.  After  punc- 
turing the  skin  he  collects  the 
blood  with  a  pipette  directly 
connected  with  the  tube  of  the 
viscosimeter. 

The  apparatus  of  W,  Hess* 
depends  upon  a  somewhat  differ- 
ent principle.  He  does  not  com- 
pare the  time  of  flow,  but  the 
distances  fluids  may  be  sucked 
up.  His  apparatus  consists  of 
two  capillaries  connected  with  a 
T-tube,  through  which  fluids  are 
sucked  with  a  rubber  bulb; 
through  one  capillary  water  is 
sucked,  and  through  the  other 
blood  or  some  other  fluid  that  is 
to  be  investigated.  From  the 
ratio  between  the  distances  to 
which  the  two  fluids  are  sucked  through  the  capillaries,  the  viscosity 
may  be  directly  determined.  The  apparatus  has  certain  special 
advantages;  the  horizontal  position  of  the  capillaries  eliminates  the 
influence  of  the  specific  gravity;  and  since  water  and  colloid  are 
simultaneously  tested,  the  errors  of  temperature  are  reduced  to  a 
minimum  and  calculations  for  correction  are  unnecessary. 


Fig.  24.    Viscosimeter,     (H.  A. 
Determan.) 


Melting,  Coagulation  and  Solidification  Temperatures. 

The  determination  of  the  melting,^  coagulation  and  solidification 
temperatures  has  the  same  significance  for  colloids  as  the  measure- 
ment of  the  melting  point  has  for  crystalloids. 

Coagulation  by  Heat.  The  fluid  to  be  investigated  is  placed  in 
a  test  tube,  in  a  water  bath.     The  contents  of  both  test  tube  and 

1  In  the  case  of  jellies  it  is  only  possible  to  speak  of  a  "  period  of  liquefaction  "; 
for  the  sake  of  simplicity  I  employ  the  expression  "melting  point." 


METHODS  OF  COLLOID  RESEARCH  115 

water  bath  must  be  stirred  and  a  thermometer  must  be  placed  in 
each.  The  test  tube  must  be  illuminated  by  a  uniform  and  pro- 
tected source  of  light.  It  is  advisable  to  make  a  number  of  prelimi- 
nary determinations  of  the  coagulation  point  before  making  the 
final  reacUng. 

Wolfgang  Pauli  distinguishes  the  following  different  appear- 
ances in  coagulation:  clear,  opalescent,  slightly  cloudy,  milky  trans- 
lucent, milky  opaque,  finely,  medium  or  coarsely  flocculent  in  slightly 
cloudy  or  clear  fluid.  These  various  aspects  are  strongly  dependent 
on  the  dilution  and  the  salt  content  of  the  solution,  and  the  latter 
has  the  greater  influence  on  the  temperature  of  coagulation. 

Melting  and  Solidification  Temperature.  To  determine  the 
melting  point  of  gelatin,  agar,  etc.,  W.  Pauli  and  P.  Rona*  used  an 
apparatus  that  is  similar  to  that  of  E.  Beckmann  for  determining 
the  freezing  point.  The  melting  point  is  the  temperature  at  which 
the  layer  surrounding  the  thermometer  melts. 

H.  Bechhold  and  J.  Ziegler*-  used  an  air  bath,  in  which  a  tube 
containing  the  jelly  is  placed  alongside  the  thermometer.  The  solid 
jelly  is  weighted  with  5  gm.  of  mercury.^  The  melting  point  is  the 
temperature  at  which  the  mercury  breaks  through  the  jelly.  Since 
it  is  difficult  to  observe  the  melting  of  the  jelly  and  the  thermometer 
at  the  same  time,  the  authors  use  an  acoustic  device  (metronome) 
which  is  described  in  the  original  papers  and  which  is  recommended 
for  similar  observations. 

Swelling. 

The  methods  of  measuring  swelling,  i.e.,  the  water  taken  up  by  a 
gel,  are  very  inexact.  The  increase  of  volume,  the  gain  in  weight  or 
the  'pressure  of  swelling  may  be  determined. 

Volume  Increase.  Equal  quantities  of  fibrin  may  be  placed  in 
test  tubes  and  covered  with  different  solutions;  we  then  observe 
how  high  the  fibrin  rises  upon  swelling  (M.  H.  Fischer,  see  p.  68, 
Fig.  7).  The  increase  in  volume  consists  of  the  decrease  in  volume 
of  the  swelling  gel  plus  the  volume  of  the  water,  so  that  the  determi- 
nation has  an  error,  inasmuch  as  the  contraction  of  the  gel  during  the 
swelling  is  unknown.  This  error  is  negligible  in  comparison  with 
the  other  experimental  errors. 

Increase  of  Weight.  This  method  introduced  by  P.  Hofmeister 
is  somewhat  more  accurate.  The  total  solids  of  the  swelling  sub- 
stance (gelatin,  muscle,  etc.)  are  determined  and  the  substance  either 
in  a  dry  or  a  swollen  state  is  placed  in  a  solution.     The  weight  deter- 

1  This  apparatus  is  made  by  C.  Gerhard,  Bonn,  Germany,  dealer  in  chemical 
utensils. 


116 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


mined  before  and  after  the  stay  in  the  solution  gives  the  amount  of 
fluid  taken  up  or  lost.  Before  weighing,  the  jelly  is  to  be  wiped 
with  filter  paper  or  a  cloth,  in  order  to  free  it  from  the  adhering 
fluid.  Fluid  is  pressed  out  in  this  method  also,  especially  in  the  case 
of  very  much  swollen  material,  by  the  weight  of  the  jelly  itself  as 
well  as  by  its  contraction  in  the  air;  this  fluid  is  dried  off  and  cannot 

be    taken    account    of    in    the 
weighing. 

Swelling  Pressure.  The  deter- 
mination of  this  factor  offers  the 
greatest  prospect  for  an  exact 
method,  especially  as  the  appa- 
ratus of  J.  Reinke*  may  be 
adapted  for  other  swelling  sub- 
stances. J.  Reinke  used  his 
apparatus  (Fig.  25)  to  measure 
the  swelling  pressure  of  laminaria 
(a  sea  weed).  The  dry  algae  are 
placed  in  the  bore  F  of  the  metal 
cylinder  M.  On  top  of  the  algse 
rests  the  piston  A,  perforated  with 
fine  holes.  From  E  water  may 
penetrate  to  the  algae  through  the 
holes.  As  the  mass  of  algae  swells 
it  raises  the  piston  and  the  rod 
ABD,  which  may  carry  various 
weights.  The  lifting  power  is 
indicated  on  the  dial.  The 
theory  of  the  apparatus,  however, 
requires  careful  investigation  to 
establish  the  relationship  be- 
tween sv/elling  pressure,  water 
absorbed  and  the  lift. 

The  apparatus  of  E.  Posnjak 
(see  Fig.  26)  offers  less  theoretical  difficulties  but  may  be  employed 
only  for  pressures  up  to  6  atmospheres.  The  principle  employed 
follows:  the  substance  to  be  investigated,  Q,  is  placed  at  the  bottom 
of  a  tube  G  which  is  closed  by  a  porous  clay  cell  T.  The  swelling 
tube  dips  into  a  vessel  of  water.  To  overcome  the  swelling  pressure 
the  whole  vessel  is  filled  with  mercury  which  is  connected  with  a 
manometer  M.  The  swelling  substances  may  be  placed  under  a 
given  pressure  by  permitting  compressed  gas  to  flow  from  a  steel  cylin- 
der g.     The  details  of  the  experiments  appear  in  the  original  paper. 


Fig.  25.     J.  Reinkes'  apparatus  for 
measuring  swelling  pressure. 


METHODS  OF  COLLOID  RESEARCH 


117 


Flocculation. 

Observations  of  the  flocculation  of  a  suspension  or  colloidal  solu- 
tion determine  whether  it  behaves  as  a  hydrophile  or  a  hydrophobe 
colloid;  furthermore,  they  show  the  electric  charge  and,  under  cer- 
tain conditions,  the  presence  of  a  protective  colloid.  The  method  of 
the  experiment  is  very  simple:    a  suspension  or  colloidal  solution  is 


'■6as 
Cylinder 


Swelling  Substance — 

Fig.  26. 


C/ay 


Thimble 


divided  among  a  large  number  of  test  tubes,  diminishing  quantities 
of  electrolytes  (NaCl,  CaCl2,  FeCls)  are  added  and  the  test  tubes  are 
filled  to  equal  volumes  with  a  solvent  (water,  physiological  salt  solu- 
tion, etc.).  After  the  test  tubes  have  been  exposed  at  uniform  tem- 
perature for  a  given  time  (1  to  24  hours),  they  are  examined  for 
flocculation. 

In  comparative  experiments  it  is  necessary  that  suspensions  or  solutions  have 
the  same  concentration.  On  account  of  the  small  amount  of  sohd  substance 
little  is  to  be  accomplished  by  determinations  of  total  sohds.  It  is  frequently  de- 
sh-able  to  prepare  a  large  quantity  of  a  standard  solution  to  last  a  long  time,  and 
frequently  the  determinations  may  be  made  colorimetrically.  I  am  accustomed 
to  prepare  mastic  suspensions  by  dropping  1  per  cent  alcoholic  mastic  solutions 
into  water  which  is  being  energetically  stirred.  The  suspension  is  filtered  through 
rather  dense  filter  paper  and  tested  for  transparency  in  a  beaker  having  parallel 
sides,  to  one  side  of  which  various  printed  fines  have  been  glued.  The  suspen- 
sion is  diluted  until,  with  a  definite  illumination,  a  certain  size  tj^je  can  just  be 
read. 


118 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


It  is  necessary  to  test  especially,  whether  true  flocculation  has  occurred,  i.e., 
whether  the  particles  have  really  gathered  in  flocks  or  whether  they  have  only 
sunk  to  the  bottom  under  the  influence  of  gravity. 

In  the  case  of  fine  measurements,  Jena  glass  that  has  been  steamed  is  used, 
because  the  leaching  out  of  aUcali  from  the  glass  may  give  rise  to  errors. 

This  method  gives  information  not  only  concerning  flocculability, 
but  is  at  the  same  time  quantitative;  it  informs  us  concerning  "thres- 
hold values"  and,  under  some  circumstances,  concerning  the  rate  of 
flocculation. 

In  order  to  work  with  very  small  quantities,  test  tubes  are  used 
which  are  narrowed  at  their  lower  ends.  If  there  is  insufficient  fluid 
even  for  this  determination  we  may  employ  a  "hanging  drop,"  in 
other  words,  drops  are  mixed  on  a  cover  glass,  and  this  is  placed  up- 
side down  on  a  slide  with  a  depression  ground  into  it,  so  that  the 
drop  will  hang.  The  cover  glass  is  ringed  with  vaseline  to  hold  it 
to  the  slide.  This  method  is  more  qualitative  and  is  especially 
adapted  for  the  agglutination  of  bacteria. 

It  is  desirable  in  all  cases  to  prepare  series  of  experiments.  It 
may  occur  as  the  result  of  "irregular  series"  that  high  and  low  con- 
centrations result  in  flocculation,  whereas  flocculation  may  not  occur 
in  medium  concentration. 


Electric  Migration. 

Electric  migration  reveals  the  nature  of  the  charge  of  a  colloid. 

The  most  primitive  arrangement  for  migration  experiments  is  a 

beaker  in  which  are  suspended 
two  platinum  electrodes  that 
are  part  of  the  circuit  of  a 
direct  current  of  at  least  60 
volts.  This  is  to  be  recom- 
mended only  for  simple  demon- 
strations in  the  lecture  room, 
where  migration  must  be  exhib- 
ited quickly.  Because  of  the 
changes  in  reaction  due  to  elec- 
trolysis, the  results  are  very  in- 
exact. It  is  preferable  to  use  a 
U-shaped   tube    or   an    arrange- 


FiG.  26a. 


m 

Simple  apparatus  for  elec- 
tric migration. 


ment  such  as  is  shown  in  Fig.  26a.  The  middle  glass  jar  contains 
the  colloid  to  be  tested,  and  is  united  by  the  U-shaped  return  bends 
filled  with  water  to  the  two  outer  beakers  which  also  contain  water 
and  into  which  dip  the  electrodes  EE.     For  research  work  I  employ 


METHODS  OF  COLLOID  RESEARCH 


119 


Fig.  27.     Bell  apparatus.     (H.  Bechhold.) 


H.  Bechhold's  "Bell  apparatus."^  The  colloid  solution  to  be  tested 
is  placed  in  the  glass  vessels  AA  (see  Fig.  27)  which  are  connected 
by  a  tube.  The  vessels  are  closed  below  by  the  membranes  MM 
(best  for  the  purpose  is  fish  bladder  or  the  like).  The  tube  R 
allows  for  the  expansion  of 
the  fluid  caused  by  the  heat 
of  the  electric  current.  The 
bell  apparatus  is  placed  in  two 
separate  glass  vessels  (crystal- 
lizing dishes)  GG;  the  mem- 
branes are  immersed  in  the 
water,  into  which  also  dip 
the  electrodes  EE.  The  advantages  of  the  apparatus  are:  the 
great  surface  of  colloid  solution;  the  products  transferred  do  not 
come  in  contact  with  the  electrodes  and  each  may  be  conven- 
iently and  separately  collected  and  ex- 
amined; the  current  must  pass  through 
the  entire  colloidal  solution;  the  ap- 
paratus may  be  easily  sterilized  and  the 
free  surfaces  may  have  toluol  layered 
over  them.  The  apparatus  does  not  get 
out  of  order  very  easily.  L.  Michaelis 
has  avoided  the  change  in  reaction  due 
to  electrolysis  at  the  electrodes  by  using 
nonpolarizable  electrodes.  Fig.  28  amply 
explains  the  apparatus.  The  electrodes, 
e.g.,  zinc  or  silver  wire,  are  dipped  into 
the  vessels  1  and  5,  which  are  filled 
with  zinc  sulphate  and  NaCl  solution 
respectively. 

Migration  experiments  are  usually  very 
difficult  to  perform.  Since  the  nature  of  the  charge  may  also  be 
determined  by  adsorption,  by  employing  positive  and  negative 
adsorbents,  this  latter  method  is  preferable,  because  it  is  simpler. 


Fig.  28.  Migration  apparatus 
with  non-polarizable  elec- 
trodes.    (L.  Michaelis.) 


Optical  Methods. 

There  is  a  certain  relation  between  the  cloudiness  (Tyndall  effect) 
of  a  fluid  and  its  content  in  suspended  particles  or  colloid.  On  this 
account  various  authors  (Kamerlingh  Onnes  and  Keesom,  Meck- 
lenburg, WiLKE  and  Handovsky)  have  constructed  instruments  to 


^  To  be  had  from  the  Vereinigten  Fabriken  für  Laboratoriumsbedarf,  BerUn  N. 
Schamhorst  Str. 


120 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


measure  the  amount  of  cloudiness  so  that  they  might  determine  from 
it,  the  content  of  dissolved  colloid  in  the  fluid. 

As  yet  they  have  not  been  applied  to  biocolloids,  and  the  relation 
between  the  clouding  of  media  in  a  fluid,  the  intensity  of  the  light  and 
cloudiness  yields  a  complicated  curve.  On  this  account  it  is  still 
impossible  to  determine  the  value  of  these  instruments  aptly  termed 

by  Mecklenburg,  tyndallmeters,  for  the 
study  of  biocolloids. 

There  is  need  of  such  an  instrument. 
[P.  A.  KoBER  has  devised  a  very  satisfac- 
tory nephelometer  which  has  found  ex- 
tensive application  in  biology,  especially 
by  Bloor.     See  Journal  of  Industrial  and 


Fig.  28a.    Kober  Nephelometer. 


Engineering  Chem.,  Vol.  VII,  p.  843.  Tr.]  I  have  always  felt  the  want 
of  being  able  to  determine  the  exact  content  of  a  bacterial  suspension 
by  some  sort  of  tyndallmeter.  Such  an  instrument  must  be  very  simple 
to  manipulate,  which  is  not  the  case  with  the  existing  instruments. 

The  colloid  content  of  a  solution  is  well  measured  for  certain 
purposes  by 

The  Fluid  Interferometer. 

The  fluid  interferometer^  was  originally  devised  to  determine  the 
concentration  or  change  in  concentration  of  crystalloid  solutions. 
According  to  Marc  it  is  also  available  for  light  or  yellowish  colloidal 
^  Made  by  Carl  Zeiss,  Jena. 


METHODS  OF  COLLOID  RESEARCH 


121 


solutions,  but  not  for  those  deeply  colored.  It  depends  on  the  follow- 
ing principle :  when  parallel  beams  of  light  pass  through  a  narrow  slit, 
as  the  result  of  refraction  a  broad  band  of  light  with  parallel  dark 
bands  (interference  bands)  is  seen  on  the  opposite  wall.  If  light  is 
permitted  to  fall  on  the  same  spot  through  a  second  parallel  slit,  the 
bands  of  light  will  interfere  and  very  fine  sharp  lines  will  be  obtained 
which  may  be  greatly  magnified.  When  a  different  medium,  that  is 
water  or  a  solution  of  salt  or  colloid  is 
placed  behind  one  slit  the  interference 
bands  move  to  one  side  depending  on 
the  refractive  index.  If  the  process  is 
reversed  by  a  set  of  glass  prisms  or 
something  similar,  it  is  possible  to  read 
on  the  adjusting  screw  of  the  appa- 
ratus the  difference  of  refractive  index. 
Fig.  28b  shows  a  cross  section 
through  the  interferometer.  A  is  the 
chamber  with  the  standard  water,  B 
the  chamber  for  the  test  solution,  C  the  window  for  viewing  the 
interference  bands.  With  dilute  solutions  the  concentration  increases 
in  proportion  to  the  scale  on  the  graduated  drum.  For  more  concen- 
trated solutions  a  standard  must  be  set  in  each  case.  Technical  de- 
tails of  the  readings  may  be  found  in  Marc's  paper  (he.  cit.).  He 
has  thus  far  used  the  interferometer  mainly  to  determine  adsorption 
and  for  studying  the  colloid  content  in  drinking  water  and  sewage. 


Fig.  28b.     Fluid  Interferometer. 


Ultramicroscopy. 

Ultramicroscopy  permits  the  recognition  of  certain  optical  in- 
homogeneities,  and  depends  upon  the  use  of  dark  field  illumination. 
Ultramicroscopes  magnifying  from  750  to  1500  diameters  serve  in 
principle  the  same  purpose  as  the  ordinary  microscope.  They  have 
the  advantage  over  the  latter  that  without  staining  or  extensive 
preparation,  even  living  objects,  spirilla,  etc.,  become  visible  to  the 
eye;  bright  on  a  dark  background.  Ultramicroscopy  with  a  one  hun- 
dred thousand  fold  magnification  has  solved  important  theoretical 
questions  of  colloid  chemistry.  By  reason  of  the  conditions  of 
light  refraction  its  value  is  chiefly  confined  to  inorganic  colloids. 

In  the  ordinary  microscope  the  field  is  usually  bright,  while  the 
object  is  more  or  less  dark  against  its  surroundings.  In  the  ultra- 
microscope,  only  the  rays  of  light  reflected  from  the  object  reach  the 
observer's  eye  and  permit  the  object  to  stand  out  bright  against  the 
dark  background. 


122 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


In  this  dark  field  illumination  the  form  of  the  objects  are  not 
given,  but  every  point  appears  as  a  small  bright  disc,  which  under 
some  circumstances  may  be  surrounded  by  one  or  several  rings  of 
light.  The  ultramicroscope  is  especially  suited  for  the  recognition  of 
inhomogeneities  in  a  medium. 

Apart  from  the  recognition  of  form,  the  field  of  application  of  the 
microscope  was  enormously  extended  by  the  invention  of  the  ultrami- 
croscope. 

At  about  seven  hundred  diameters'  magnification,  the  limit  of  the 
available  microscopical  magnification  is  reached  theoretically  and 
practically,  i.e.,  revelation  of  new  details  ceases.  The  ability  to  make 
particles  visible  in  the  ultramicroscope  is  almost  unlimited,  provided 
only  a  sufficiently  strong  source  of  light  is  available.  Practically, 
the  limits  of  visibility  in  our  latitude  with  the  best  sunlight  is  about 
10  MM  (1  MM  =  1  miUionth  part  of  a  millimeter).  [Zsigmondy  gives 
5  MM.     Tr.j 

For  our  purposes,  two  types  must  be  distinguished:  (a)  Ultra- 
microscopes  for  the  study  of  colloids.  They  permit  the  observation 
of  objects  or  inhomogeneities  down  to  10  mm  and  require  very  bright 
sources  of  light  —  sunlight  reflected  from  a  heliostat,  or  electric  arc 
lights.  (&)  Ultramicroscopes  for  the  study  of  organized  materials 
(microorganisms,  animal  and  plant  cells),  suitable  for  the  study  of 
objects  no  smaller  than  0.1  m-  Welsbach  or  Nernst  lights  in  com- 
bination with  suitable  lenses  furnish  sufficient  illumination. 

Ultramicroscopy  for  the  Study  of  Colloidal  Solutions. 

The  original  slit-ultramicroscope  constructed  by  H.  Siedentopf 
and  R.  Zsigmondy  with  rectangular  arrangement  of  the  optical  axes 


Fig.  31a.     Slit-ultramicroscope. 


is  nowadays  only  employed  for  the  study  of  solid  objects  (glasses) 
and  on  this  account  may  be  omitted  from  consideration  in  bio- 
colloid  investigations.  Recently  Zsigmondy  has  adapted  the  original 
slit-ultramicroscope  to  immersion.     A  large  proportion  of  the  light 


METHODS  OF  COLLOID  RESEARCH 


123 


rays  in  the  path  of  the  object  examined  are  lost  by  refraction.  Very 
small  objects,  such  as  bacteria  are  too  faintly  illuminated  to  be 
visible  by  his  "dry  system."  On  this  account,  a  highly  refractive 
fluid  (water  or  cedar  oil)  is  placed  between  the  object  and  the  objec- 


FiG.  29.     Illumination  of  the  cardioid  ultramicroscope. 


tive  (of  wide  aperture),  which  permits  many  more  rays  to  pass  from 
the  object  into  the  objective.  It  was  impossible  to  use  immersion 
in  the  earlier  sht-ultramicroscope  because  the  illuminating  (ßi)  and 


\/^/^^/^^/^/^//^^^^^////////////////A 


ft  *  M 

Fig.  30.  Course  of  the  Hght  rays 
through  the  cardioid  condenser. 
(H.  Siedentopf.) 


Fig.  31.     Quartz  chamber  for  the 
cardioid  ultramicroscope. 


the  examining  objective  (Ss)  could  not  be  brought  sufficiently  close 
together  (see  Fig.  31a).  This  difficulty  was  overcome  through  an 
improved  method  of  construction  by  the  optical  works  of  R.  Winkel 
of  GoTTiNGEN  (see  Fig.  30).  A  drop  of  the  fluid  to  be  examined  is 
placed  between  the  two  immersion  objectives  of  wide  aperture  or 


124 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


they  dip  into  a  small  trough  containing  the  fluid  for  examination. 
The  illumination  intensity  of  the  "immersion  ultramicroscope "  is 
much  greater  than  the  original,  and  particles  are  made  visible  which 
formerly  had  quite  eluded  observation;  the  contrast  effect  in  the 
intensely  dark  field  is  quite  perfect.  For  biologists,  the  ultramicro- 
scope with  a  cardioid  condenser  is  at  present  the  most  important 
instrument.  It  permits  the  use  of  twenty  times  as  much  Hght  as  the 
slit-ultramicroscope,  "practically  the  maximum  available  from  the 
source  of  light." 

The  construction  of  the  apparatus  is  shown  in  Fig,  29.     c  con- 
tains an  electric  arc  lamp  with  a  perforated  sleeve  cap  d  to  cut  out 


Fig.  32.     Holder  for  the  quartz  chamber  em- 
ployed with  the  cardioid  ultramicroscope. 


Fig.  33.     Flasks  for  storing 
ultrawater.      (A.  Haak.) 


interfering  light.  An  illuminating  lens  e  passes  the  light  sharply 
downward,  through  a  glass  trough  filled  with  water,  to  the  center  of 
the  microscope  mirror. 

The  water  trough  serves  to  remove  the  heat  rays  or  when  neces- 
sary acts  as  a  color  filter.  The  microscope  mirror  throws  the  light 
perpendicularly  through  the  cardioid  condenser,  which  replaces  the 
Abbe  condenser  in  the  microscope.  It  is  evident  from  the  diagram 
of  the  cardioid  condenser  (Fig.  30)  that  the  various  ascending  rays 
strike  the  slide  e  obliquely  by  reason  of  the  double  reflection  from  the 
two  spherical  surfaces  and  that  thus,  all  the  light  is  utilized  for 
illumination;  only  the  rays  reflected  by  the  object  take  the  usual 
path  through  the  objective  and  the  ocular  to  the  observer's  eye. 
Water  is  used  for  immersion. 

With  the  cardioid  ultramicroscope  the  object  is  placed  between 
slide  and  cover  glass  as  in  ordinary  microscopy.  For  reasons  we 
shall  revert  to  later,  a  slide  with  a  special  quartz  chamber  (Fig.  31) 
is  used,  which  is  held  in  the  holder  (Fig.  32). 


METHODS  OF  COLLOID  RESEARCH 


125 


There  are  a  number  of  precatitions  to  be  observed  in  working  with  the  ultra- 
microscope.  Since  every  impurity  makes  a  point  of  light  in  the  field,  it  is  neces- 
sary to  employ  optically  clear  water.  Such  water  is  prepared  according  to  R. 
ZsiGMONDY  by  distillation  through  a  silver  condensing  tube,  or  according  to  H. 
Bechhold  by  ultrafiltration  through  a  very  tight  ultrafilter  (6  to  10  per  cent). 


Fig.  34.     Dark-field  illumination  for 
the  examination  of  organisms. 


For  collecting  and  storing,  only  Jena  glass  vessels  should  be  employed.  Ground 
glass  stoppers  or  corks  are  to  be  avoided  because  they  always  yield  fine  dust. 
I  have  found  the  suggestion  of  W.  Biltz  serviceable ;  he  coated  the  stoppers  with 


Fig.  34a. 


tin  foil.     I  recommend  a  storage  flask  for  ultrawater  manufactured  by  A.  Haak 
in  Jena  (Fig.  33). 

Neither  water  nor  alcohol  should  show  microscopically  the  slightest  Faraday- 
Tyndall  effect,  but  wherever  illuminated,  only  a  very  faint  shimmer,  whitish 
in  the  case  of  water  (ultrawater),  bluish  in  the  case  of  alcohol  (ultra-alcohol). 


126 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Though  a  skilled  ultramicroscopist  usually  recognizes  impurities  from  irregular 
intensity  of  illumination  and  color  of  the  submicrons,  as  well  as  by  differences  in 
motion,  the  most  extreme  care  is  necessary  in  ultramicroscopic  work. 

Cover  glasses  for  the  upper  chamber  should  be  of  quartz,  3/4  mm.  thick. 
The  usual  methods  of  cleaning  (cloths,  brushes,  elderpith  and  Japanese  tissue) 
are  to  be  avoided,  as  particles,  which  may  cause  much  trouble,  are  broken  off; 
scratches,  tears,  and  impurities  arising  from  dry  cleaning  increase  the  adsorption 
of  colloids  on  the  chamber  walls  and  reveal  their  own  ultramicroscopic  pictures 
independently  of  the  coUoid  particles.  The  chamber  and  cover  slip  must  always 
be  cleaned  in  the  following  fashion:  nothing  is  touched  by  hand,  only  forceps 
with  platinum  points  or  with  a  loop  of  platinum  wire  about  them  may  be  used. 
The  apparatus  is  placed  in  a  hot  mixture  of  concentrated  H2SO4  and  sodium  bi- 
chromate, then  washed  with  tap  water  and  finally  conductivity  water.  The  water 
must  be  removed  with  ultra-alcohol  and  finally  collodion  is  poured  over  the  cleaned 
surface.  Before  use,  the  collodion  skin  may  be  easily  raised  at  one  corner  and 
removed. 

On  forcing  the  chamber  and  cover  slip  in  the  holder,  it  is  necessary  to  avoid 
screwing  too  tight  or  tensions  will  arise  which  gradually  equalize  themselves  and 
cause  striations  which  are  very  disturbing. 


Ultramicroscopes  for  the  Study  of  Organized  Material. 

The  apparatus  for  this  purpose  may  be  adjusted  to  any  microscope. 
Special  preparation  of  the  objects  is  unnecessary. 

Objects  difficult  to  make  visible  by  staining  or  which  are  too  small 
to  see  alive  with  the  ordinary  microscope  are  especially  suitable  for 


y^ N 

k 

II 

1 

Fig.  35.    Abbe  condenser  with 
central  opacity. 


Fig.  36.  Paraboloid  condenser  for  the 
dark-field  illumination  of  organisms. 
(From  H.  Siedentopf.) 


this  method  of  investigation,  e.g.,  spirilla,  protoplasmic  structures, 
etc.  A  picture  is  obtained  similar  to  that  with  Burri's  India  ink 
method,  in  which  the  rest  of  the  field  is  blackened  with  India  ink, 
while  the  objects  appear  bright.  Oblique  illumination  reveals  in- 
homogeneities  and  structures  which  would  be  invisible  even  with 


METHODS  OF  COLLOID  RESEARCH  127 

staining.  In  the  description  of  the  investigations  of  N.  Gaidukow, 
E.  RÄHLMANN,  etc.,  we  shall  return  to  this  topic.  The  optical  system 
depends  on  the  fact  that  the  central  rays  of  light  reflected  from  the 
mirror  of  the  microscope  are  cut  out  by  a  disc,  whereas  the  lateral 
rays  which  strike  the  object  obliquely  are  utilized. 

The  simplest  and  cheapest  arrangement  is  the  one  by  which  a 
central  blind  is  placed  in  the  diaphragm  carrier  of  the  Abbe  illumi- 
nating apparatus  (see  Fig.  35),  yet  this  arrangement  is  not  recom- 
mended on  account  of  the  faint  illumination  and  the  difficulty  in 
centering. 

Much  to  be  preferred,  because  of  the  strong  illumination,  is  Sieden- 
topf's  paraboloid  condenser  (see  Fig.  36).  It  is  adapted  to  the  study 
of  living  bacteria  and  especially  for  thin  organized  structures. 

The  thicker  the  preparation  the  weaker  must  be  the  objective. 

Preparations  must  be  made  with  greater  cleanliness  than  for  bright  field 
illumination,  though  such  scrupulous  care  is  not  necessary  as  for  the  cardioid 
condenser. 

The  sUde  must  have  a  definite  thickness  (not  less  than  1.1  mm.  or  more  than 
1.4  mm.). 

The  object  to  be  studied  is  placed  on  a  sUde  moistened  with  a  drop  of  physio- 
logical salt  solution  and  a  cover  glass  adjusted  so  that  there  are  no  bubbles.  The 
water  pressed  out  at  the  sides  is  absorbed  and  the  rim  is  sealed  tight  with  wax 
(1  part  wax,  2  parts  rosin).  A  drop  of  water  without  bubbles  is  placed  between 
the  slide  and  the  condenser.  Neither  water  nor  oil  is  used  between  the  shde 
and  the  objective  (dry  system). 

[  Other  ultramicroscopes  have  been  devised  by  Cotton  and  Mouton,  and  by 
IvANOWSKi  (made  by  E.  Leitz).  See  also  L' Ultra-microscope  by  Paul  Gaston, 
Paris,  1910.    Tr.] 


An  asterisk  (*)  after  an  author's  name  refers  to  a  reference  in  the  index  of 
names. 

PART   II. 
THE  BIOCOLLOIDS. 

With  the  exception  of  water,  inorganic  salts  and  a  few  organic 
substances  as,  for  instance,  urea  and  sugar,  only  colloids  exist  in 
plant  and  animal  organisms,  and  if  we  except  water,  the  colloids 
quantitatively  far  exceed  the  crystalloids.  This  appears  reasonable 
when  we  consider  the  respective  roles  of  crystalloids  and  colloids  in 
the  organism.  We  may  compare  living  organisms  to  a  city,  in  which 
the  colloids  are  the  houses  and  the  crystalloids  are  the  people  who 
traverse  the  streets,  disappearing  into  and  emerging  from  the  houses, 
or  who  are  engaged  in  demolishing  or  erecting  buildings.  The  colloids 
are  the  stable  part  of  the  organism:  the  crystalloids  the  mobile  part, 
which  penetrating  everywhere  may  bring  weal  or  woe.  Because  they 
have  only  a  transitory  use,  we  find  in  the  organism  only  a  small 
number  and  a  small  quantity  of  organic  crystalloids.  In  plants  we 
encounter  the  most  important  organic  crystalloid,  sugar,  on  its  way 
from  its  place  of  origin  to  the  place  where  it  is  used,  or  in  depots, 
such  as  buds,  roots,  fruits,  etc.,  where  it  is  either  changed  into  an 
insoluble  form  of  carbohydrate,  into  starches  and  related  products, 
or  its  retreat  is  cut  off  by  the  drying  of  the  stem  from  which  the 
fruit  depends.  In  its  course  we  may  tap  great  quantities  of  sugar, 
as  in  the  birch,  maple  and  palm  when  they  are  "in  sap."  If  for 
any  reason  it  becomes  mobilized  again  in  the  depots,  large  quantities 
of  sugar  may  be  formed.  In  wild  plants  the  amount  of  sugar  is 
rarely  very  great;  it  is  otherwise  with  cultivated  plants  where  as  the 
result  of  cultivation  sugar  is  stored  with  no  advantage  to  the  plant, 
e.g.,  sugar  beets,  sugar  cane  and  common  beets.  At  times  a  certain 
biological  purpose  may  be  associated  with  sugar  formation,  e.g.,  the 
sugar  formation  in  fruits  for  the  purpose  of  their  dissemination. 
The  fruit  is  always  the  biological  object  and  serves  to  perpetuate 
the  species,  not  the  individual.  The  development  of  a  greater 
quantity  of  crystalloid  as  sugar  in  fruit  is  therefore  not  surprising, 
since  the  fruit  has  completed  its  service  for  the  individual  plant. 
Elsewhere,  we  find  the  carbohydrates  only  in  colloidal  and  most 
often  even  in  insoluble  form.     I  refer  to  starches,  cellulose  and  gums. 

129 


130  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Like  plants,  the  animal  organism  has  the  power  of  changing  carbohy- 
drates into  crystalloids.  Ferments  change  the  starches  into  sugar, 
in  fact  cellulose  which  is  so  resistant  to  chemical  attack  is  made 
soluble  in  the  intestine  of  vegetarians,  so  that  it  can  enter  the  animal 
body.  As  soon  as  the  crystalloid  forms  of  carbohydrate  have  passed 
the  intestinal  wall  they  are  transferred  to  the  main  depot,  the  liver, 
where  they  remain  in  the  stable  colloidal  condition  as  animal  starch, 
glycogen.  We  also  find  glycogen  in  most  of  the  other  organs,  where- 
as the  mobile  state  of  carbohydrate,  grape  sugar,  occurs  only  in 
minimal  quantities  (0.08  to  0.12  per  cent),  in  fact  only  just  so  much 
as  is  necessary  for  the  production  of  energy. 

Fats,  too,  are  found  in  the  truly  soluble  form  (e.g.,  soaps)  in  plants, 
only  in  the  germs  of  seeds;  and  in  animals,  probably  only  at  the 
moment  when  they  pass  through  the  wall  of  the  intestines.  They  have 
hardly  passed  the  intestine  when  they  immediately  regain  their  colloidal 
condition  of  emulsion,  and  are  carried  in  that  condition  to  their  depots. 

The  same  statements  hold  for  proteins.  Crystalloid  cleavage  prod- 
ucts are  found  in  germinating  seeds  and  in  minimal  quantities  in  the 
vascular  paths;  in  plants,  asparagin;  in  animals,  among  others, 
urea,  uric  acid  and  ammonia  salts.  The  organism  strives  its  utmost 
to  retain  the  colloidal  condition.  Hardly  have  the  crystalloid 
cleavage  products  of  albumin  which  have  been  formed  in  the  stomach 
and  intestines  passed  through  the  intestinal  wall,  than  they  are 
straightway  changed  back  into  the  colloidal  form,  so  that  their  re- 
turn may  be  cut  off.  The  crystalloid  combustion  products  are  given 
an  avenue  of  escape  through  the  kidneys. 

Physiological  chemistry  deals  with  the  role  of  the  carbohydrates, 
fats  and  proteins  apart  from  water  and  the  inorganic  salts.  In  the 
study  of  biocolloids,  water  and  salts  cannot  be  neglected,  because 
water  and  salts  are  an  indispensable  part  of  the  colloids;  no  colloid 
can  exist  in  the  organism  without  them,  because  they  condition  the 
turgescence  which  is  characteristic  of  living  colloid. 

In  the  case  of  cells  with  true  membranes,  salts  may  determine  at 
times  the  balance  of  osmotic  pressure  within  and  without  the  cell. 
This  general  fact  does  not  explain  the  necessity  of  the  various 
kinds  of  anions  and  cations  (K,  Na,  Ca,  Mg,  CI,  SO4,  PO4,  CO2); 
the  balance  in  osmotic  pressure  may  be  maintained  by  any  non- 
electrolyte  (e.g.,  sugar)  and  yet  a  cell  cannot  be  kept  alive  in  an  iso- 
tonic sugar  solution.  Inorganic  salts  have  specific  relations  to  certain 
organs  to  which  we  shall  refer  later;  they  are  the  expression  of 
characteristic  sharply  defined  physical  states  assumed  in  the  presence 
of  given  quantities  of  water  and  salt,  by  the  proteins,  carbohydrates, 
etc.,  of  which  the  organs  consist. 


THE  BIOCOLLOIDS  131 

.J 

Chemistry  in  general,  and  physiological  chemistry  in  particular, 
aims  to  investigate  the  structure  of  individual  chemical  substances, 
and  thus  explain  their  properties  by  splitting  them,  synthesizing 
them,  and  comparing  the  regenerated  (rearticulated)  substance  with 
the  original,  to  see  if  it  is  the  same  or  different.  Unfortunately,  so 
far  as  the  colloidal  constituents  of  the  organism  are  concerned,  they 
are  still  far  from  their  goal,  especially  in  the  case  of  carbohydrates 
and  proteins.  Here  colloid  chemistry  enters  and  attempts  to  com- 
prehend and  where  possible  to  regulate  the  behavior  of  the  finished 
product.  Colloid  chemistry  is  not  occupied  with  the  parts  of  the 
machine,  but  with  the  machine  itself.  The  chemist  splits  the  pro- 
teins into  Polypeptids,  amino-acids,  etc.,  but  the  student  of  biocolloids 
avoids  such  profound  attacks  and  strives  to  keep  the  molecule  in- 
tact so  far  as  possible,  studying  its  outward  form,  the  chemical 
points  of  attack  offered  by  the  unmutilated  molecule,  its  behavior  to 
changes  which  may  occur  under  normal  and  pathological  conditions, 
as  well  as  those  brought  about  by  drugs. 

I  wish  here  to  emphasize  one  other  point.  Only  a  few  substances 
occur  in  the  organism  that  are  suitable  for  study  by  the  physiological 
chemist.  Serum  albumin  and  globulin,  the  starches  and  some  of  the 
fats,  are  unquestionably  substances  which  may  be  separated  from  the 
organism  without  losing  some  of  their  essential  properties,  but  they 
are  exceptions.  The  substances  usually  studied  by  physiological 
chemists  are  those  which  have  already  suffered  considerable  modi- 
fication. The  organism  possesses  neither  glue,  histone  nor  myosin, 
and  even  if  we  knew  the  exact  chemical  constitution  of  glue,  this 
would  throw  no  light  upon  the  properties  and  the  function  of  cartilage 
and  the  fibrils  of  connective  tissue  from  which  it  is  derived.  But 
even  without  knowing  the  chemical  composition  of  glue,  I  believe 
that  it  would  be  possible,  with  the  methods  of  colloid  chemistry  alone, 
to  collect  a  series  of  observations  which  would  afford  valuable  con- 
clusions concerning  the  chemical  mechanisms  of  such  tissues. 

A  time  will  come  when  the  old  physiological  chemistry  and  the 
new  chemistry  of  the  biocolloids  will  meet  and  the  two  opposite 
ends  of  the  tunnel  shall  be  united.  We  shall  first  try  to  learn  the 
properties  of  the  intact  colloid  molecule  of  the  colloid  particle. 
The  following  chapters  on  carbohydrates,  lipoids  and  proteins  should 
be  read,  bearing  this  statement  in  mind. 


CHAPTER  VIII. 
CARBOHYDRATES. 

As  the  name  indicates,  we  classify  as  carbohydrates  a  group  of 
substances  containing  carbon  and  the  elements  of  water,  i.e.,  0  and 
H  in  the  proportion  of  1  :  2. 

We  owe  our  knowledge  of  the  constitution  of  the  lower  members 
of  this  group,  the  crystalloid  water-soluble  sugars,  largely  to  the  in- 
vestigations of  Emil  Fischer.  The  same  difficulties  which  we 
encounter  in  the  study  of  all  colloidal  substances  interfere  with  de- 
termining the  constitution  of  the  higher  colloidal  members  of  this 
group,  the  saccharocolloids.  There  is  at  present  no  means  of  positively 
recognizing  the  purity  and  the  individuality  of  the  substance  studied 
or  its  derivatives.  It  is  true  '^that  we  may  crystallize  individual 
colloidal  carbohydrates,  e.g.,  inulin,  which  as  a  rule  naturally  occurs 
in  crystals,  but  all  we  have  said  on  page  71  concerning  the  crystalli- 
zation of  colloids  in  general  applies  to  inulin. 

Because  of  their  common  occurrence,  the  most  important  sac- 
charocolloids are  the  starches,  vegetable  and  animal  (glycogen),  and 
also  cellulose.  Next  in  importance  come  the  various  gums  and  pec- 
tinous  plant  juices.  Dextrins  which  are  also  usually  colloidal  are 
really  cleavage  products  of  the  starches. 

A  host  of  individual  facts  have  been  derived  from  the  enormously 
extensive  utilization  of  starches,  as  food,  cereals  and  potatoes,  for 
fermented  liquors,  beer  and  brandy,  as  sizing,  etc.,  and  of  cellulose 
(in  the  textile  industries  and  paper  manufacture).  It  is  only  recently 
that  there  has  been  manifested  an  effort  to  reach  a  general  view- 
point such  as  colloid  science  has  made  possible.     (E.  Fouard.*) 

Starch,  obtained  from  starchy  grains,  is  an  amorphous  white 
powder  which  migrates  in  the  electric  current  to  the  anode,  it  exhibits 
an  acid  character  chemically,  since  it  adsorbs  dissolved  alkalis  (with 
the  exception  of  NH4OH)  and  hydroxids  of  the  heavy  metals, 
probably  thus  forming  amylates.  It  does  not  adsorb  acids  or  salts. 
(A.  Rakowski.*)  Since  phosphoric  acid  is  always  present  in  native 
starches  and  in  the  diastatic  cleavage  of  phosphorus-containing 
dextrins,  we  may  assume  with  M.  Samec  that  there  is  a  carbohydrate 
phosphoric  acid  complex  probably  an  ester  (amylophosphate) . 
Starch  has  a  great  reversible  swelling  capacity  in  water  (pore  swelling). 

133 


134 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


In  swelling  there  is  a  great  loss  of  volume,  i.e.,  the  volume  of  the 
swollen  starch  is  less  than  that  of  the  dry  starch  plus  the  water 
necessary  for  swelling,  as  was  shown  in  exhaustive  experiments  by 
H.  RoDEWALD.*  This  contraction  is  about  8  per  cent,  when  20  per 
cent  of  water  is  taken  up.  Swelling  is  accompanied  by  the  liberation 
of  heat,  which,  according  to  E.  Wiedemann  and  Charles  Lude- 
KiNG,*  amounts  to  about  6.6  calories  per  gram.  H.  Rodewald 
studied  the  phenomenon  more  thoroughly  and  found  a  diminution 
in  the  amount  of  heat  liberated  with  increasing  water  content.  The 
following  is  an  abbreviated  table  of  his  results: 


Per  cent  of  watet 

contained  in  100  gm. 

dry  starch. 

Approximate  per  cent 

of  heat  Hberated  per 

gm.  dry  starch. 

0.23 

28.11 

2.39 

22.60 

4.58 

18.19 

9.59 

10.28 

18.43 

3.54 

If  we  add  more  water  to  starch,  and  heat  to  55°-70°  C,  by  "solu- 
tion swelling,"  we  get  a  jelly-like  mass,  starch  paste,  which  dissolves 
on  continued  heating  in  more  water.  This  solution  coagulates 
when  it  is  frozen.  G.  Malfitano  and  A.  N.  Moschkoff*  utilize 
this  property  of  starch  solution  to  obtain  a  starch  free  from  mineral 
substances.  Demineralized  starch  on  being  mixed  with  suitable 
salts  shows  all  the  properties  of  the  different  forms  of  starch.  These 
investigators  are  therefore  of  the  opinion  that  the  various  modifica- 
tions in  the  properties  of  the  natural  starch  granules  are  due  to  mineral 
admixtures. 

E.  FouARD,*  by  means  of  acids,  freed  starches  from  their  inor- 
ganic elements  and  obtained  a  substance  which  formed  an  unstable 
colloidal  solution  in  water.  Heat,  alkalis  and  alkaline  salts  made  the 
solution  more  permanent,  whereas  cold,  acids  and  acid  salts  favored 
jelly  formation.  On  ultrafiltering  his  starch  solutions,  E.  Fouard 
found  that  in  accordance  with  their  concentration,  a  given  fraction 
of  the  solution  always  passed  through  collodion  membranes.  He 
concluded  from  this,  that  for  every  concentration  of  the  starch  solu- 
tion a  balance  exists  between  the  coarser  particles  and  the  molec- 
ularly  dissolved  (hydrolyzed?)  starches.  Unfortunately,  the  work 
of  E.  Fouard  contains  no  information  relative  to  the  permeability 
of  the  collodion  membranes,  so  that  it  is  impossible  to  arrive  at  any 
conclusion  in  reference  to  the  size  of  the  suspended  and  the  dissolved 
starch  particles. 


CARBOHYDRATES  135 

On  account  of  their  great  surface  development,  the  adsorptive 
capacity  of  starches  is  very  great.  As  has  been  said,  when  they 
swell  they  adsorb  water,  dyes,  etc.  A  very  characteristic  adsorption 
compound  is  formed  with  iodin.  lodin  is  the  best  known  reagent  for 
starches;  by  it  they  are  stained  blue.  It  was  formerly  beheved 
that  iodin  and  starch  united  chemically;  W.  Biltz  showed  that  it  is 
merely  an  adsorption.  According  to  the  degree  of  dispersion,  iodin 
solution  is  blue,  red,  orange  or  yellow,  inasmuch  as  the  starch  solu- 
tion acts  as  a  protective  colloid  (W.  Harrison*).  There  are,  in 
addition,  varieties  of  starch  which  give  at  once  a  brownish  red  or  a 
wine  red  color  with  iodin.  Inulin  and  lichenin  are  colored  yellow 
by  iodin. 

The  swelling  and  pasting  of  starches,  hydration,  is  analogous  to  the 
swelling  of  proteins,  which  is  a  preliminary  to  their  hydrolytic  cleavage. 
The  swelling  of  starches  is  favored  by  electrolytes,  especially  alkalis, 
so  that  swelling  commences  at  a  much  lower  temperature  in  their 
presence.  For  this  purpose  the  anions  are  especially  important  and 
in  fact,  in  a  lyotropic  series,  similar  to  that  of  acid  albumin.  See 
page  152  (M,  Samec).* 

Starch  paste  increases  the  surface  tension  of  water  (Zlobicki*). 
A  solution  of  starch  in  water,  as  well  as  one  of  dextrin,  dissolves  less 
CO2  than  pure  water  (according  to  A.  Findlay*).  (A  gelatin  solu- 
tion dissolves  more  CO2  than  pure  water!) 

'Under  the  influence  of  dilute  acids  or  diastatic  ferments,  the  starch 
molecule  takes  up  water  and,  step  by  step,  breaks  into  small  frag- 
ments, soluble  starches,  amylodextrin,  various  dextrins  some  of  which 
crystallize,  and  finally  into  grape  sugar.  The  larger  the  fragments 
the  more  marked  is  their  colloidal  character. 

As  the  result  of  osmometric  experiments  W.  Biltz  *  arrived  at  the 
following  molecular  weights: 

Amylodextrin 22,200-20,500 

Higher  achroodextrin 11,700-  8,200 

Erythrodextrin 6,800-  3,000 

Acid  dextrin 4,000 

Lower  achroodextrin 1,800-  1,200 

Dextrin  (C6Hio06)6 905 

Commercial  dextrin 6,200-  2,700 

"Soluble  starch"  (according  to  H.  Friedenthal*^  produces  a 
definite  lowering  of  the  freezing  point,  which  is  proportionate  to  the 
amount  of  the  substance  that  is  dissolved. 

CrystalUzable  dextrins  [amyloses  (C6Hio05)6]  prepared  by  H 
Pringsheim*  and  Eissler  combine  with  iodin  to  form  iodin-addition 
compounds  which  dissolve  like  iodin  starches  in  cold  water  with  a 
transitory  blue  color. 


136  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Commercial  dextrins  which  are  mixtures  of  starch  fragments  of 
different  size  are  almost  entirely  held  back  by  impermeable  ultra- 
filters  (10  per  cent)  (H.  Bechhold*^). 

Closely  related  to  the  starches  is  inulin,  the  reserve  carbohydrate 
in  dahha  bulbs  and  the  roots  of  Inula  helenium,  etc.,  as  well  as 
lichenin,  which  occurs  in  many  lichens,  especially  Iceland  moss. 
Unlike  the  starches,  inulin  and  lichenin  are  soluble  in  water  without 
forming  a  paste  and  form  yellow  adsorption  compounds  with  iodin 
(see  p.  135). 

Besides  these,  a  series  of  starches  have  been  identified,  some  of 
which  show  differences  in  their  final  cleavage  products,  the  sugars. 
As  yet  they  have  not  been  studied  colloid-chemically. 

In  its  biological  function,  animal  starch,  glycogen,  resembles  the 
plant  starches  closely,  and  in  its  colloid  properties  stands  midway 
between  these  and  inulin.  It  swells  in  cold  water  and  forms  with 
it  an  opalescent  hydrosol.  The  electric  current  carries  it  to  the 
anode  (Z.  Gatin-Gruszewska*).  With  iodin  it  forms  according  to 
its  concentration,  a  brownish  yellow  to  deep  red  adsorption  com- 
pound. 

The  internal  friction  of  glycogen  solutions  have  been  studied  by 
F.  BoTTAZzi  and  G.  D'Errico*  as  well  as  by  J.  Friedländer.* 

Glycogen  is  split  up  by  acids  and  ferments,  and  according  to  the 
degree  of  hydrolysis  we  find  all  sorts  of  fragments,  from  the  highly 
colloidal  to  the  easily  diffusible  grape  sugar.  E.  Rählmann*^  fol- 
lowed this  process  with  the  ultramicroscope. 

The  glucosides  must  be  mentioned  in  this  connection.  They  are 
compounds  of  the  aliphatic  and  the  aromatic  series  with  sugars, 
which  may  be  split  into  their  components  by  acids  or  ferments.  In 
the  vegetable  kingdom  they  include  very  active  pharmacologic  and 
toxic  substances,  such  as  digitalis  glucoside,  phloridzin  and  saponins. 
Recently  several  glucosides  have  been  discovered  in  the  animal  organ- 
ism, e.g.,  cerebron  in  the  human  brain.  Though  some  glucosides, 
e.g.,  amygdalin  and  myronic  acid  are  unquestionably  crystalloids, 
others,  e.g.,  saponin,  are  entirely  colloidal.  Since  we  know  very  little 
of  the  biological  significance  of  glucosides,  it  is  evident  that  we  do  not 
know  what  importance  may  be  ascribed  to  the  crystalloid  form  in 
one  and  the  colloidal  form  in  the  other. 

The  gums  are  carbohydrates  which  are  widely  distributed  through- 
out the  vegetable  kingdom.  Some  of  them  play  a  part,  in  many 
respects  analogous  to  that  of  fibrin  in  the  animal  kingdom,  since  they 
solidify  on  issuing  from  a  wound,  thus  sealing  it.  Best  known  of 
the  gums  are  gum  arabic,  carraghen  and  cherry  gum,  while  agar,  de- 
rived from  Japanese  sea  weed,  is  of  especial  importance  in  bacteri- 


CARBOHYDRATES  137 

ology.     Finally,  we  must  mention  the  pectinous  plan-t  juices,  which 
unlike  the  true  gums  are  slightly  or  not  at  all  soluble  in  water. 

The  gums  are  typical  examples  of  hydrophile  colloids;  they  swell 
into  jellies  in  water,  and  on  adding  more  water  pass,  at  an  indefinite 
point,  into  solution.  Rise  of  temperature  shifts  this  point  in  favor  of 
solution,  though  it  is  by  no  means  immaterial  at  what  condition  of 
swelling  the  heating  occurs.  If,  for  instance,  agar  has  been  allowed  to 
swell  in  cold  water  for  a  long  time,  it  immediately  becomes  a  homo- 
geneous solution  on  warming.  If  solid  agar  is  heated  in  water,  we 
get  a  lumpy  suspension  of  agar  in  water,  which  only  very  gradually 
becomes  a  homogeneous  sol.  It  is  evidently  necessary  for  each 
particle  of  agar  to  have  the  amount  of  water  necessary  for  solution 
in  close  proximity  before  it  is  warmed;  otherwise  the  swelling  will 
occur  but  slowly  from  the  outside,  where  there  is  an  excess  of  water, 
and  proceed  inward,  since  the  peripheral  particles  of  agar  hold  the 
water  until  they  are  dissolved.  Indeed,  the  phenomenon  is  one  which 
depends  on  the  size  of  the  surface;  the  large  mass  with  relatively 
small  surface  dissolves  more  slowly  than  the  same  mass  divided, 
i.e.,  with  a  relatively  increased  surface.  Solutions  of  gum  do  not 
dialyze.  In  my  opinion  little  attention  need  be  paid  to  the  determi- 
nation of  their  osmotic  pressure,  since  traces  of  electrolytes  which 
cannot  be  removed,  suffice  to  simulate  it.  I  know  of  no  studies 
on  the  electrical  migration  or  on  the  diffusion  coefläcients  of  gums. 
[W.  M.  Bayliss  has  recently  determined  the  viscosity  and  osmotic 
pressure  against  water  and  Ringers'  solution  of  gum  acacia,  gelatin 
and  amylopectin.  He  recommends  the  use  of  gum  and  gelatin  in 
saline  infusions  as  a  method  of  maintaining  blood  pressure.  The 
more  prolonged  action  of  such  infusions  he  attributes  to  the  osmotic 
pressure  of  the  colloids.  Proceedings  of  the  Royal  Society  of  London, 
Series  B,  No.  89,  pp.  380-393.     Tr.] 

Gums  usually  diminish  the  surface  tension  of  water.  The  tr  of  a 
20  per  cent  solution  of  gum  arable  is  9  per  cent  lower,  and  a  dilute 
solution  of  agar  5  per  cent  lower  than  that  of  water  (G.  Quincke). 
Some  kinds  of  gum  increase  the  surface  tension  of  water  (Zlobicki*). 

The  general  facts,  stated  on  page  66,  hold  for  the  swelling  and 
shrinking  of  gums.  On  sweUing,  the  heat  hberated,  according  to 
E.  Wiedemann  and  Chas.  Ludeking,*  is  9.0  cal.  per  gm.  for  gum 
arable  and  10.3  cal.  per  gm.  for  gum  tragacanth.  Wo.  Pauli*^  found 
that  a  rise  of  temperature  accompanied  the  sweUing  of  carraghen. 

The  significance  of  crystalloids  for  swelling  and  turgor  has  been 
studied  chiefly  in  gelatin.  In  the  case  of  the  gums,  other  than  agar, 
no  investigations  of  this  point  have  been  made.  Though  the  prob- 
ability of  many  similarities  exists,  an  absolute  parallelism  cannot  be 


138  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

assumed.  Thus,  for  instance,  the  melting  point  of  gelatin  is  raised 
by  grape  sugar  and  glycerin,  whereas  that  of  agar  is  reduced.  NaCl 
elevates  the  melting  point  of  agar  and  depresses  that  of  gelatin  (H. 
Bechhold  and  J.  Ziegler*^). 

Agar  has  a  very  strong  tendency  to  gelatinize;  even  1  gm.  per 
liter  gelatinizes  at  0°.  This  great  gelatinizing  capacity  led  Robert 
Koch  to  make  his  culture  media  of  agar,  and  permitted  him  to  grow 
cultures  of  bacteria  on  solid  media  at  body  temperature.  Gelatin 
media  which  had  been  used  at  first  melt  at  37°  C,  and  could,  ac- 
cordingly, only  be  used  at  room  temperature. 

Electrolytes  as  well  as  nonelectrolytes  alter  the  gelatinization 
time  of  agar.  Nitrates,  iodids,  sulphocyanids,  benzoates,  urea  and 
thiourea  lengthen  it;  chlorids,  bromids,  acetates  and  salts  of  poly- 
basic  acids  shorten  it. 

Cellulose  is  for  plants  what  bones  are  for  animals.  It  forms  the 
framework  which  maintains  their  shape.  If  it  is  to  fulfill  this  function 
it  must  be  insensitive  to  the  chemical  influences  of  the  plant  juices, 
and  must  not  be  able  to  swell.  Wooden  relics  are  by  no  means  un- 
common; only  in  exceptional  instances  are  fats  and  proteins  or 
gelatinous  constituents  seen  after  thousands  of  years,  and  then  only 
under  very  unusually  favorable  conditions,  as  in  the  desert  climate 
of  Egypt.  Wood,  even  uncarbonized,  is  a  common  relic  not  only  for 
Egyptian  archaeologists  and  travellers  in  the  Turanian  deserts,  but 
it  has  frequently  been  preserved  in  our  own  climate  and  even  in 
water.  Oak  bridge  piles  dating  from  Roman  times  have  been  found 
in  the  Rhine,  wood  carvings  and  wooden  buckets  in  the  springs  of 
Salzburg,  fragments  of  boats  of  the  lake  dwellers,  in  the  Swiss  lakes, 
and  those  of  the  Vikings  in  the  peat  bogs  of  North  Germany  and 
Jutland.  Stability  of  form,  in  other  words,  a  slight  swelling  capacity, 
makes  wood,  next  to  stone,  metal  and  bone,  suitable  for  many  pur- 
poses. 

Cellulose,  the  principal  constituent  of  wood,  is  extremely  inactive 
and  is  only  split  up  into  soluble  sugars  (chiefly  grape  sugar)  by 
strong  chemical  action  (acids  concentrated  or  under  pressure),  or 
by  specific  ferments  (bacteria  in  the  intestines  of  ruminants). 

Cellulose  not  only  has  a  high  adsorptive  capacity  for  dyestuffs, 
but  even  true  suspensions  are  fixed  at  its  surface.  For  this  reason 
cellulose  has  recently  been  used  like  charcoal  a's  a  clarifier  and  as  a 
filter  for  turbid  liquids. 


CHAPTER  IX. 
LIPOIDS. 

''  Lipoids  "  is  the  collective  name  for  fatty  substances.^  Many  of 
them  are  not  moistened  by  water;  however,  this  property  is  not 
characteristic  of  all  lipoids. 

Fats  and  oils  are  esters  of  higher  fatty  acids,  usually  with  glycerin, 
which  may  be  substituted  by  other  higher  alcohols;  for  instance,  a 
palmitic  acid  ester  of  cetyl  alcohol  occurs  in  spermaceti,  found  in  the 
skull  of  the  sperm-whale.  Though  in  other  fats  all  three  hydroxyls 
of  glycerin  are  replaced  by  fatty  acid  radicals,  in  the  lecithins  only 
two  fatty  acid  radicals  occur,  and  the  third  hydroxyl  group  is  re- 
placed by  a  phosphoric  acid-cholin  radical.  Cholin  is  a  trimethyl- 
oxy  ethylammoniumhydroxid . 

Formula  of  Fats.  Formula  of  Lecithin. 

CHzO-fatty  acid  CHaO-cholin-phosphate 

CHO-fatty  acid  CHO-fatty  acid 

CHsO-fatty  acid  CHaO-fatty  acid 

Finally,  we  must  consider  Cholesterins  and  isocholesterins,  which  we 
may  regard  as  complex  terpenes. 

The  characteristic  fats,  the  triglycerides,  are  universally  distributed 
in  the  animal  body,  where  they  play  an  important  part  in  maintaining 
the  body  heat,  while  in  plants  they  are  of  much  less  importance. 
Lecithins  are  found  distributed  throughout  the  animal  organism,  not 
only  in  the  chief  depots,  the  brain,  nervojiis  tissue  generally  and  the 
egg  yolk,  but  in  every  cell,  every  organ,  even  in  the  lymph,  blood 
corpuscles  and  muscles.  In  plants  too,  lecithin  is  widely  distributed, 
occurring  in  the  seeds. 

The  fact  that  lecithins  occur  in  all  parts  of  the  body  is  an  evidence 
of  their  great  biological  importance.  So  far  as  may  be  gathered 
from  previous  researches,  they  play  an  important  role  in  the  life- 

1  Various  investigators  give  different  definitions  of  the  term  "  lipoids."  Bang 
uses  it  most  inclusively  and  regards  everything  in  the  body  soluble  in  organic 
eolvents  as  lipoid;  S.  Loewe  gives  it  the  narrowest  scope,  and  includes  only  sub- 
stances which  form  colloid  solutions  in  organic  solvents  {e.g.,  cephalin,  cerebrosid). 

139 


140  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

processes  of  the  cells  and  in  the  adjustment  of  the  metabolism  be- 
tween cells  and  their  surrounding  media.  The  same  is  true  of 
Cholesterin  which  is  frequently  associated  with  them. 

Fats  and  oils  are  not  soluble  in  water  and  aqueous  solutions;  but 
instead  they  are  easily  emulsified  by  a  great  variety  of  substances. 
A  few  drops  of  lye  suffice  to  make  the  finest  sort  of  subdivision  of 
oil  in  water.  It  is  still  an  open  question,  whether  this  is  accomplished 
by  the  lye  itself,  or  whether  it  is  due  primarily  to  soaps,  which  are 
formed  from  the  free  fatty  acids  always  present  in  fats  and  oils,  and 
which  themselves  act  as  emulsifiers.  Soluble  soaps,  i.e.,  the  fatty 
acid  salts  of  the  alkalis,  possess  remarkable  fat-emulsifying  proper- 
ties; this  property  is  also  shared  by  the  intestinal  juice,  the  pan- 
creatic juice  and  the  bile.  Emulsions  of  fat  and  oil  usually  occur 
in  alkahne  solution,  while  on  the  other  hand  acids  produce  floccula- 
tion.  There  are  exceptions  to  this,  e.g.,  the  lipase  of  the  castor 
bean  emulsifies  fat  in  acid  solution,  and  milk  curded  by  rennet  yields 
a  stable  acid  emulsion  on  digestion  in  pepsin-hydrochloric  acid.  In 
general,  fat  emulsions  behave  like  hydrophile  colloids;  they  are  not  as 
easily  coagulated  by  neutral  salts  as  are  hydrophobe  colloids  or  other 
suspensions. 

Milk  is  a  natural  emulsion  of  fat  (see  p.  345  et  seq.). 

Though  in  the  examples  given  so  far,  fat  has  been  the  dispersed 
phase  and  water  or  the  aqueous  solution  the  dispersing  medium, 
conversely,  water  and  aqueous  solutions  may  be  incorporated  in  fats. 
In  this  case  fat  is  the  dispersing  medium  and  the  aqueous  solution 
the  dispersed  phase.  Instances  of  this  condition  are  butter,  cold 
cream,  which  is  cooling  because  of  the  water  it  contains,  lanolin,  as 
well  as  many  salves  and  liniments.  Structures  like  cream  and  whipped 
cream  occupy  a  characteristic  intermediate  position. 

Lecithin  behaves  in  a  very  peculiar  way.  It  forms  an  emulsion 
with  water  of  its  own  accord;  indeed  like  a  protein  it  swells  up  in 
water  into  a  turbid  colloidal  solution,  without  dissolving.  It  may 
be  said  that  it  occupies  a  place,  in  respect  to  its  colloidal  properties, 
between  the  emulsifiable  fats  and  the  hydrophile  colloids,  closely  ap- 
proaching the  latter. 

0.  Forges  and  E.  Neubauer*  studied  its  properties  by  experi- 
menting upon  the  coagulation  of  lecithin  emulsions. 

The  precipitating  action  of  neutral  salts  is  in  a  lyotropic  series 
similar  to  that  for  acid  albumin,  in  which  the  greatest  effect  is  pro- 
duced by  the  anions.  Salts  of  the  alkaline  earths  and  the  heavy 
metals  frequently  yield  "zones  of  inhibition"  as  described  on  page 

84.     It  is  remarkable  that  neither  HgCU  nor  Hg(CN)2  even  in  -p 


LIPOIDS  141 

jf 
Concentration  cause  precipitation.     This  is  in  thorough  accord  with 
the  solubility  of  such  substances  in  fats. 

Lecithin  acts  towards  colloids  and  suspensions  (ferric  hydroxid, 
mastic  suspension)  like  any  other  colloid  which  migrates  to  the 
anode.  Similarly  charged  colloids  cause  no  precipitation  (and 
lecithin  may  even  act  as  a  protective  colloid  for  mastic) ;  oppositely 
charged  colloids  produce  flocculation  in  suitable  mixtures  (ferric- 
oxid  hydrosol).     Saponin  clears  lecithin  suspensions. 

Alcoholic  lecithin  solutions  are  much  more  stable  in  the  presence  of 
salts  than  aqueous  solutions.  Mercuric  chlorid  is  an  exception. 
Alcoholic  lecithin  solutions  protect  some  other  colloids,  e.g.,  albumoses, 
from  the  precipitating  action  of  alcohol.     (L.  Michaelis  and  P. 

IlONA*^.) 

Ethereal  lecithin  solutions  cause  some  otherwise  insoluble  sub- 
stance to  dissolve  in  ether  (e.g.,  NaCl  and  grape  sugar).  This  prop- 
erty is  evidently  due  to  the  fact  that  in  ethereal  solution,  lecithin 
has  a  great  capacity  for  taking  up  water. 

Cholesterin,  according  to  the  investigations  of  0.  Porges  and  E. 
Neubauer,*  is  a  hydrophobe  colloid.  Its  aqueous  emulsion  behaves 
like  a  mastic  suspension  in  the  presence  of  a  large  variety  of  salts. 
The  same  is  true  for  its  behavior  with  other  colloids.  In  neutral 
solution  it  is  precipitated  by  certain  proportions  of  albumin  and 
saponin.  Lecithin  may  act  as  a  protective  colloid  for  Cholesterin. 
Cholesterin  forms  a  true  solution  in  alcohol  and  ether,  and  in  such 
solutions  exhibits  no  colloid  precipitation  reactions. 


CHAPTER  X. 
PROTEINS. 

We  designate  as  proteins  a  group  of  nitrogenous  colloids  which 
are  the  chief  constituents  of  animals  and  plants.  They  consist  en- 
tirely or  chiefly  of  substances  which  contain  quantitatively: 

Per  cent. 

C 50-55 

H 6.5-7.3 

N 15-17.6 

0 19-24 

S 0.3-  2.4 

One  of  the  chief  characteristics  of  most  of  the  dissolved  albumins 
is  their  coagulability  when  heated.  The  effect  of  heat  on  undis- 
solved proteins  is  shown  by  the  loss  of  their  capacity  to  swell;  they 
are  "denatured."     Hydrophile  colloids  become  hydrophobe. 

A  host  of  the  most  diverse  substances  are  included  under  the 
generic  term  "albumin."  It  includes  water-soluble  substances  such 
as  egg  and  serum  albumin,  and  substances  soluble  in  saline  solutions, 
as  globulin,  vitellin,  myosin  and,  finally,  such  substances  as  are  solu- 
ble neither  in  aqueous  nor  in  saline  solution,  for  example,  fibrin. 
We  know  that  there  exists  in  each  plant  and  in  each  animal  a  distinct 
serum  albumin  and  a  distinct  serum  globulin,  etc.  In  the  chapter  on 
"Immunity  Reactions,"  we  shall  return  to  the  species-native  charac- 
teristics (Artspezifität)  of  proteins  (see  p.  194).  We  shall  not  speak 
of  these  distinctions  here,  but  we  shall  dwell,  rather,  upon  the  prop- 
erties that  the  different  proteins  possess  in  common. 

Colloid  research,  in  a  negative  way,  by  destroying  a  large  number 
of  false  conceptions,  has  been  of  great  service  to  the  chemistry  of 
proteins;  and  it  is  in  a  position  to  establish  new  principles,  since 
only  a  few  proteins  crystallize  and,  with  others,  common  methods  of 
purification  are  unavailable.  Absolutely  misleading  methods  have 
been  relied  upon  to  separate  and  distinguish  proteins.  It  was  form- 
erly believed,  e.g.,  that  the  coagulation  temperature  of  different  proteins 
varied,  but  colloid  investigations  demonstrated  that  small  quantities 
of  electrolytes  could  raise  or  depress  it  to  a  great  extent.  By  pre- 
cipitation with  copper  sulphate,  E.  Harnack  believed  that  he  had 
obtained  characteristic  copper  albuminates,  and  other  observers  that 
they  had  obtained  characteristic  silver  or  calcium  albuminates. 
Colloid  chemistry  has  shown  that  the  different  amounts  of  copper, 

142 


PROTEINS  143 

silver,  etc.,  contained  in  such  precipitates  depend  upon  the  concen- 
trations of  the  solutions  of  albumin  and  of  electrolyte,  and  that 
precipitates  of  constant  constitution  are  always  obtained  under  the 
same  conditions.  Fr.  N.  Schulz  and  R.  Zsigmondy  showed  that 
crystallized  egg  albumin  which  had  adsorbed  colloidal  metallic  gold, 
recrystallized  with  it. 

As  the  result  of  such  observations  we  become  very  sceptical  con- 
cerning the  ''purity"  of  proteins.  However,  it  is  just  such  explana- 
tion of  earUer  errors  which  shows  us  upon  what  facts  we  may  really 
depend,  and  gives  to  science  a  new  method  and,  in  part,  a  new  course. 

Before  we  describe  the  few  proteins  which  have  been  studied  colloid- 
chemically,  we  shall  consider  briefly  some  of  their  general  properties. 

One  of  the  most  characteristic  properties  of  many  proteins  is 
coagulation.  It  may  be  brought  about,  either  by  a  rise  of  temper- 
ature (heat  coagulation)  or  by  chemical  means. 

Most  of  the  coagulations  due  to  the  salts  of  the  light  metals  and 
some  of  those  due  to  the  alkaline  earths  are  reversible,  i.e.,  the 
coagulations  reverse  themselves  by  the  addition  of  more  water.  Heat 
coagulation  and  coagulation  due  to  many  of  the  salts  of  the  heavy  metals 
are  irreversible.  The  coagulations  due  to  alcohol,  acetone  and  ether 
are  intermediate,  that  is,  the  coagulation  produced  is  at  first  soluble 
in  water  but  becomes  insoluble  after  a  while.  Globulin  which  has 
been  preserved  for  a  time  in  pure  water  behaves  in  a  similar  way, 
for  it  then  becomes  less  soluble  in  salt  solutions. 

Though  reversible  coagulation  may  be  viewed  as  a  purely  physical 
salting  out  (see  under  this  heading)  a  chemical  change  must  be  assumed 
in  the  cases  of  irreversible  coagulation.  Many  heavy  metals  form 
insoluble  complexes  with  albumin  (see  p.  157).^  Irreversible  coagu- 
lation by  heat,  alcohol,  etc.,  may  be  explained,  possibly,  by  a  chemical 
transformation.  The  fact  that  the  H  ion  concentration  diminishes 
after  heat  coagulation  is  in  favor  of  this  view  (Sörensen  and  Jurg- 
SEN,*  H.  Chick  and  C.  J.  Martin,*  Guaglieriello*)  .  In  the  case 
of  heat  coagulation,  water  appears  to  enter  the  albumin  molecule, 
because  absolutely  dry  hemoglobin  and  egg  albumin  may  be  heated 
to  120°  C.  without  losing  their  solubiHty  in  water  (H.  Chick  and 
C.  J.  Martin*).  Possibly  this  is  the  initial  stage  of  hydrolysis,  since 
according  to  Berczeller*  the  surface  tension  of  salt-poor  albumin 
solutions  is  temporarily  depressed  upon  boiUng,  just  as  occurs  upon 
hydrolysis  by  pepsin,  trypsin,  etc.  Irreversibly  coagulated  albumi- 
nous pellicles  may  be  formed  merely  by  shaking  with  air  (see  p.  34). 

1  [Sansum  has  shown  that  after  the  absorptionof  alethaldoseof  4mg.  perkilo 
of  mercuric  chlorid,  no  treatment  avails.  Jour.  Am.  Med.  Association,  vol.  70, 
p.  824.    Tr.] 


144  COLLOIDS  IN  BIOLOGY,  AND  MEDICINE 

Though  native  albumins  are  usually  hydrophile,  they  become  hydro- 
phobe upon  heat  coagulation.  Trace's  of  acids  and  salts  cause  precipita- 
tion.    The  precipitate  of  albumin  induced  by  freezing  is  irreversible. 

Albumin  may  be  partly  changed  to  globuhns,  and  ultimately 
coagulated  and  precipitated  by  light,  particularly  light  of  short  wave 
length  (G.  Dreyer  and  Hausen,  Chalupecky),  ultraviolet  rays  are 
particularly  intense  in  their  action  (Bovie).  This  is  especially 
significant  for  some  future  explanation  of  the  action  of  sunlight  on  the 
organism.  Schanz  attributes  to  it  the  clouding  of  the  crystalline 
lens  in  cataract.  The  rays  of  shortest  wave  length,  the  Roentgen 
rays,  coagulate  albumin. 

A  number  of  proteins  have  been  crystallized  (e.g.,  egg  albumin, 
horse  serum  albumin,  hemoglobin,  aleuron)  and  though  the  shape 
of  the  crystal  is  characteristic  for  the  kind  of  albumin,  nevertheless 
it  is  impossible  to  obtain  the  crystals  absolutely  chemically  pure  as  in 
the  case  of  crystalloids  (see  p.  71). 

Albumin  solutions  have  been  studied  ultramicroscopically  by  E. 
Rählmann,*^  E.  von  Behring,  H.  Much,  Römer  and  C.  Siebert,* 
by  L.  Michaelis,*^  L.  Pinkussohn*  and  J.  Lemanissier.*  The 
results  expected  at  the  outset  were  not  realized,  so  that,  in  recent 
years,  there  has  been  little  heard  on  the  subject.  In  my  opinion 
this  is  unfortunate;  I  am  inclined  to  believe  that  valuable  data 
might  be  gleaned  from  a  properly  controlled  ultramicroscopic  study 
of  proteins.  It  is  evident  that  a  large  part  of  albumin  solutions  is 
amicroscopic,  so  that  only  such  portions  are  seen  as  show  a  different 
refraction  than  water  or  physiological  salt  solution.  An  albuminous 
solution  shows  a  different  number  of  ultramicrons,  entirely  depend- 
ing upon  whether  it  has  been  prepared  in  water  or  in  physiological 
salt  solution  (Michaelis);  and  with  different  dilutions  depending 
upon  the  salt  content,  a  different  number  of  small  particles  become 
visible  (Rählmann).  On  this  account  L.  Michaelis  and  J.  Lema- 
nissier do  not  share  the  opinion  of  E.  Rählmann  and  the  school  of 
E.  VON  Behring  as  to  the  suitability  of  ultramicroscopic  observa- 
tion for  the  quantitative  determination  of  albumin,  e.g.,  in  the  urine. 
Great  interest  must  attach  to  ultramicroscopic  observations  of  the 
cleavage  of  albumin  by  pepsin,^  the  influence  of  therapeutically 
active  substances  (ferric  chlorid,  alum,  tannic  acid,  silver  nitrate, 
copper  sulphate,  collargol,  etc.),  as  well  as  the  effect  of  dyes  on  solu- 
tion of  albumin  (Rählmann).  A  few  submicrons  were  found  by  J. 
Lemanissier  in  albumin  solution  and  many  in  hemoglobin,  but  they 
disappeared  in  24  hours. 

Ultrafiltration  of  albumin  solution  is  still  in  its  infancy.  H. 
Bechhold  has  shown  that  the  particles  of  serum  albumin  are  some- 

1  [Already  observed  by  J.  Alexander.  Jour.  Am.  Chem.  Soc,  Vol.  XXXII, 
p.  680,  et  seq.    Tr.] 


PROTEINS  145 

what  smaller  than  those  of  hemoglobin.  Unlike  ferments,  proteins 
are  not  strongly  adsorbed  by  filter  material. 

All  albumins  are  amphoteric  electrolytes,  i.e.,  they  yield  H  and  OH 
ions;  otherwise  expressed,  they  have  at  the  same  time  the  character 
of  weak  acids  and  of  weak  bases,  with  the  acid  character  more  or  less 
in  excess.  The  consequences  resulting  in  the  case  of  albumin  have 
been  discussed  more  extensively  on  p.  154. 

The  isoelectric  point  is  that  where  the  sum  of  the  H  and  OH  ions 
is  least.  This  point  acquired  especial  significance  from  the  studies 
of  L.  Michaelis  who  showed  that  the  isoelectric  point  was  charac- 
teristic for  each  albumin.  That  albumins  are  most  easily  precipitated 
at  this  point  was  also  demonstrated  [by  Hardy.  Tr.].  In  this  respect 
they  behave  like  crystalloid  electrolytes.  Neutral  molecules  are 
much  more  difficult  to  dissolve  than  their  ions.  Acids  sUghtly  dis- 
sociated electrically,  e.g.,  uric  acid,  salicylic  acid,  quinine,  are  much 
more  difficult  to  dissolve  than  their  strongly  dissociated  salts. 

Adsorption  phenomena  are  of  great  importance.  Proteins  may  be 
strongly  adsorbed  or,  on  the  other  hand,  exert  a  powerful  adsorption. 
The  purely  physical  phenomena  are  complicated  by  the  intermingling 
of  specific  chemical  properties  and  thus  very  decided  differences  be- 
tween the  various  groups  of  albumins  are  brought  to  light. 

Proteins  as  Adsorbed  Substances.  Adsorption  has  been  most 
carefully  studied  in  the  case  of  albumin.  As  a  result  of  its  faint 
acidity  it  is  completely  adsorbed  by  ferric  oxid  hydrogel,  but  mastic 
and  kaolin  suspensions  on  the  contrary  adsorb  it  only  in  faintly 
acid  solution  (L.  Michaelis  and  P.  Rona*).  On  this  account,  any 
suspension  may  be  employed  to  remove  albumin  from  acid  solutions, 
e.g.,  urine,  whereas  an  electropositive  adsorbent  {e.g.,  ferric  oxid  gel) 
must  be  chosen  in  the  case  of  neutral  fluids.  Although  the  distri- 
bution between  solvent  and  adsorbent  has  the  shape  of  an  adsorp- 
tion curve,  it  must  nevertheless  be  emphasized  that  the  process 
(adsorption  by  iron-oxid,  cellulose  and  kaolin)  is  only  incompletely 
reversible,  thus  resembling  the  phenomena  of  dyeing  (W.  Biltz*^). 
The  adsorption  of  euglobulin  by  kaoHn  (K.  Landsteiner  and 
Uhlirz*)  is  to  be  explained  in  a  similar  way. 

Proteins  as  Adsorbents.  Proteins  are  frequently  used  as  adsorb- 
ent both  in  a  solid  and  in  a  denatured  condition.  They  take  up 
acids,  alkahs,  salts,  dyes,  etc.,  from  solution,  in  accordance  with  the 
formula  of  an  adsorption  curve.  In  my  opinion  it  is  best  to  regard 
the  compound  as  an  adsorption  whenever  the  chemical  constitution  of 
the  adsorbed  substance  is  unknown  or  when  it,  itself,  possesses  col- 
loid properties.  To  view  the  facts  from  the  standpoint  of  chemical 
constitution  (see  p.  154),  a  viewpoint  which  presupposes  a  more 


146  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

exact  knowledge  of  the  mechanism  of  the  reaction,  seems  to  me  to  be 
a  still  more  advanced  step. 

Adsorption  by  protein  in  solution  is  more  important  than  ad- 
sorption by  solid  proteins.  By  ultrafiltration  it  might  be  possible  to 
investigate  the  distribution  between  a  dissolved  colloid  and  a  crystal- 
loid. In  this  connection  I  am  acquainted  only  with  the  investiga- 
tions of  H.  Bechhold  on  the  distribution  of  methylene  blue  between 
water  and  serum  albumin  (see  p.  26). 

Thomas  Geaham  and  R.  O.  Herzog*^  determined  the  coefficient  of 

D  cm^. 

diffusion  of  egg  albumin  and  ovomucoid   to  be    f-   •  10^.     Its 

"^  seconds 

values  are  in  the  case  of 

Egg  albumin 0.063  (at  13°  C.)  measured  by  Graham,  calculated  by 

Stefan. 

Egg  albumin 0.054  (at  15.3°  C.)  according  to  Herzog. 

Egg  albumin 0.046  (at    7.75°  C.)  according  to  Herzog. 

Egg  albumin   [crystallized  with   3.6%.  ..  .0.081    (at    16°  C.)    according  to 
Dabrowski*  (NH4)2S04]. 

Ovomucoid 0.034  (at    7.75°  C.)  according  to  Herzog. 

Glucose 

(for  comparison)  .   0.57   (at   18°  C.) 

From  these  figures  the  radius  r  of  albumin  particles  has  been 
calculated  for 

Salt-free  egg  albumin 2,43  mm 

Crystallized  egg  albumin 1,37  ßß 

[with  3.6%  (NH4)2S04] 

This  diminution  in  the  size  of  the  albumin  particles  in  the  presence  of 
(NH4)2S04  coincides  with  what  we  shall  learn  of  the  other  effects  of 
neutral  salts  on  albumin  (see  p.  151). 

When  solid  albumins  go  into  solution  there  occurs  a  diminution 
in  volume  amounting  to  about  5-8  per  cent,  as  is  the  case  with 
starches.     (H,  Chick  and  C.  J.  Martin.*) 

Egg  albumin  and  serum  albumin,  globulin,  casein  and  fibrin  have 
been  most  carefully  studied  colloid  chemically. 

ALBUMINS. 

Albumins  are  soluble  in  water,  and  in  dilute  neutral  salt,  and  in 
acid  and  in  alkaline  solutions.  They  are  usually  found  in  the  com- 
pany of  globulins  and  there  are  reasons  for  beheving  that  they  may 
be  converted  into  globulins  by  moderate  heating.  Albumins  occur 
almost  exclusively  in  serum,  in  eggs  and  in  milk;  the  existence  of 
plant  albumins  is  not  yet  definitely  established. 


PROTEINS  147 

In  the  organism  proteins  only  occur  accompanied  by  electrolytes 
which  greatly  modify  their  properties.  On  this  account  we  shall  try 
to  get  an  idea  of  albumin  unassociated  with  electrolytes  in  order  to  un- 
derstand the  influence  of  the  addition  of  electrolytes. 

Electrolyte-Free  Albumin.^ 

Wolfgang  Pauli  obtained  an  albumin  free  from  electrolytes  by 
dialysing  ox  serum  in  closed  vessels  for  eight  weeks.  After  standing 
undisturbed  for  several  weeks,  the  serum  was  filtered  and  was  foimd 
to  furnish  a  stable  crystal-clear  fluid.  Boiling  and  the  addition  of 
alcohol  completely  coagulated  the  solution.  Such  albumin  is  am- 
photeric with  a  weakly  electronegative  charge;  so  that  it  consists 
chiefly  of  neutral  and  very  slightly  ionized  particles  ^  which  migrate 
to  both  electrodes  in  an  electric  field  (L.  Michaelis*^).  According 
to  L.  Michaelis  and  P.  Rona,  the  isoelectric  point  for  serum  al- 
bumin, at  which  there  is  the  greatest  tendency  to  precipitation,  occurs 
with  an  H-ion  concentration  of  2.10~^;  for  boiled,  denatured  serum 
albumin  when  the  H-ion  concentration  is  4.10~^.  It  increases  the 
internal  friction  of  water  considerably.  If  the  friction  coefficient 
of  water  is  represented  by  1000,  a  1  per  cent  amphoteric  albumin 
solution  at  the  same  temperature  will  be  1068.  An  equimolecular 
1  per  cent  salt  solution  causes  no  demonstrable  change  in  the  coeffi- 
cient of  friction  of  water. 

Solubility  in  Albumin  Sols. 

We  shall  see  in  the  following  pages  that  albumin  usually  has  a 
powerful  influence  on  the  solubility  of  substances.  It  is  a  remark- 
able fact  that  the  solubility  of  carbonic  acid  is  the  same  in  an 
albumin  sol  as  it  is  in  water  (A.  Findlay*).  This  is  all  the  more  re- 
markable since  starches  and  gelatins,  in  contradistinction  to  albumin, 
are  very  active  in  affecting  the  solubiHty  of  carbonic  acid.  This  is 
physiologically  important,  since  serum  consequently  plays  no  part 

1  The  colloid-chemical  study  of  proteins  was  inaugurated  by  F.  Hofmeister 
and  his  pupils;  in  recent  times  they  have  been  studied  chiefly  by  Wo.  Pauli 
and  his  co-workers  in  numerous  experimental  investigations.  We  wish  espe- 
cially to  call  attention  to  Wo.  Pauli  and  H.  Handovsky,  Hofmeister's  Beitr.  z. 
Chem.  Physiol,  u.  Pathol.,  11,  415-448;  Biochem.  Zeitschr.,  18,  pp.  340-371.  Loc. 
cit.,  24,  239-262.  Further  references  in  the  text  book  of  H.  Freundlich  and  Wo. 
OsTWALD  as  well  as  in  H.  Handovsky,  KoU.-Zeitschr.,  4  and  5  (1910). 

^  Since  an  absolutely  ash-free  albumin  cannot  be  prepared  bj^  dialysis  as 
shown  by  the  investigations  of  Pauli  and  the  unpublished  experiments  of  H. 
Bechhold  and  J.  Ziegler,  it  is  apparent  that  the  question  of  the  electric  charge 
of  pure  albumin  is  not  yet  definitely  determined. 


148 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


in  respiration.  I  have  no  knowledge  of  researches  as  to  whether  the 
H-ion  concentration  of  water  containing  CO2  is  affected  by  ash-free 
albumin. 

Wolfgang  Pauli  and  M.  Samec*  have  commenced  exhaustive 
studies  into  the  influence  of  albumins  on  the  solubility  of  electro- 
lytes. They  employed  a  serum  albumin  solution  which  had  been 
dialyzed  eight  weeks  and  contained  2.23  per  cent  of  albumin.  All  the 
readily  soluble  electrolytes  investigated  showed  a  slight  decrease  in 
solubility  as  compared  with  pure  water.  The  solubilities  were  as 
follows : 


Ammonium  chlorid 

Magnesium  chlorid 

Ammonium  suphocyanate 


In  100  gm.  serum 
solution. 


27.9 

35.51 

62.06 


Contrariwise,  the  solubility  of  difficultly  soluble  electrolytes  was  de- 
cidedly increased  by  the  presence  of  albumin. 
The  solubilities  were  as  follows: 


Calcium  sulphate 

Calcium  phosphate  Ca3(P04)2 

Calcium  carbonate 

Silicic  acid 

Uric  acid r 


In  100  gm.  water. 


0.223 
0.011 
0.004 
0.023 
0.040 


In  100  gm.  serum 
solution. 


0.226 
0.021 
0.023 
0.030 
0.057 


Having  in  view  the  deposition  of  urates  in  gout,  H.  Bechhold  and 
J.  ZiEGLER*^  undertook  exhaustive  studies  of  the  solubility  of  uric 
acid  and  urates  in  electrolyte-free  serum.  Since  even  traces  of 
NaHCOs  (in  the  case  of  uric  acid)  and  Na  salts  (in  the  case  of 
Na-urate)  may  greatly  influence  the  solubility,  before  dialysing  the 
serum,  HCl  was  added  until  the  NaHCOs  was  completely  neutra- 
lizedj  and  the  last  traces  of  Na  salts  were  removed  by  repeated 
additions  of  KCl.  Each  addition  was  followed  by  dialysis.  In 
this  way  the  following  solubilities  of  Na-urate  and  uric  acid  were 
obtained  in  electrolyte-free  serum  albumin  solution  containing  7.6 
per  cent  albumin  (expressing  the  percentage  in  relation  to  the  entire 
quantity  of  protein  in  defibrinated  blood  serum)  at  37°  C. 

In  1000  gm.  serum  albumin  solution  (in  1000  gm.  water): 

Uric  acid,  549  to  668  mg 64. 9  mg. 

Monosodium  urate,  476  to  568  mg 1200  to  1500  mg. 


PROTEINS 


149 


The  ability  to  decrease  the  solubility  of  easily  soluble  electrolytes 
and  to  increase  the  solubility  of  difficultly  soluble  electrolytes  is  not 
a  specific  property  of  albumins  but  is  common  to  colloids  in  general, 
e.g.,  gelatin. 

Albumin  and  Hydrosols.  The  exhaustive  studies  of  U.  Friede- 
mann* show  that  electrolyte-free  serum  and  egg  albumin  are  pre- 
cipitated both  by  positive  and  negative  inorganic  hydrosols.  An 
optimum  precipitation  zone  exists  here,  as  it  does  in  other  colloid 
precipitations.  Excess  of  albumin  or  inorganic  hydrosol  hinders  the 
precipitation.  Addition  of  NaCl  shifts  the  zone  of  precipitation 
without,  however,  conforming  to  any  definite  law.  As  an  example 
I  might  mention  the  precipitation  (XXX)  of  albumin  by  diminish- 
ing quantities  of  molybdic  acid,  with  and  without  added  NaCl. 


Molybdic  acid. 


0.5 

0.25 

0.1 

0.05 

0.025 

0.001 

0.005 


Albumin  about 
3  per  cent. 


Degree  of  precipitation  in 
salt-free  solution. 


XXX 
0 
0 
0 

XXX 
XXX 

xx-xxx 


Degree  of  precipitation  on 

the  addition  of  4  drops  of 

10  per  cent  NaCl. 


XXX 
XXX 
XXX 
XXX 

0 

0 

0 


As  the  result  of  cataphoretic  experiments,  U.  Friedemann  be- 
lieves that  the  charge  of  proteins  towards  water  is  not  determinative 
of  their  precipitation  by  inorganic  hydrosols.  Albumin  which 
travels  to  the  anode,  notwithstanding  this  fact  gives  heavy  pre- 
cipitates with  inorganic  hydrosols  (arsenic  trisulphid,  silicic  acid, 
molybdic  acid).  There  is  much  to  justify  the  assumption  of  U. 
Friedemann  that  a  given  hydrosol,  according  to  its  charge,  collects 
at  the  +  or  —  charge  of  the  amphoteric  albumin,  thus  permitting 
its  aggregation  to  larger  complexes. 

The  albumins  appear  to  act  with  proteins  of  definite  basic  (histones) 
or  acid  character  just  as  they  do  with  inorganic  hydrosols  (U.  Friede- 
mann and  H.  F.riedenthal*). 


Influence  of  Electrolytes. 

If  an  electrolyte  is  added  to  an  amphoteric  albumin,  the  properties 
of  the  albumin  undergo  considerable  modification.  Salts,  even  in 
very  small  quantities  (hundredth  normal),  raise  the  coagulation  tem- 
perature, as  is  shown  in  the  subsequent  coagulation  temperatures 
taken  from  a  table  compiled  by  Wo.  Pauli  and  H.  Handovskt.*^ 


150 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Salt 

0 

0.01  n 

0.02  n 

0.03  n 

0.04« 

0.05« 

NaSCN 

Na2S04 

NaCl 

NaCaHsOa 

KSCN 

60.3° C. 
60.3° C. 
60.3° C. 
60.3° C. 
64.6° C. 

68 

66.7 

63.16 

66.9 

68.3 

69.7 
68 

65.7 
69.2 

70.6 
68.5 
66.4 
70.6 
69.5 

71.6 
69.1 
67.2 
71.5 

72.5 
69.7 
67.9 
72.1 
70.3 

This  table  shows  the  remarkable  fact  that  the  first  traces  of  salt 
have  a  much  greater  influence  than  somewhat  greater  concentra- 
tions. 0.01  normal  Na2S04  added  to  salt-free  albumin  raises  the 
coagulation  temperature  about  6.4°  C.  while  a  similar  addition  to 
albumin  already  containing  0.04  normal  Na2S04  raises  the  coagula- 
tion temperature  only  0.6°  C.  We  shall  show  the  significance  of  this 
fact  later. 

If  the  salt  is  more  concentrated,  coagulation  by  heat  varies;  the 
coagulation  temperature  rises  continuously  in  the  presence  of  K,  Na 
and  NH4.     Thus  for 

3  normal  KCl,  coagulation  occurs  at  75 . 6°  C. 
3  normal  NaCl,  coagulation  occurs  at  73.6°  C. 
3  normal  MgCl2,  coagulation  occurs  at  75 . 4°  C. 

The  coagulation  temperature  reaches  a  maximum  at  a  certain 
salt  concentration  and  then  falls  again  in  the  case  of  other  salts, 
especially  alkaline  earths  and  the  allied  lithium. 

Maximum  coagulation  temperature,"  C. 

6  normal  NH4CI 72.8 

2  normal  (NH4)2S04 74.3 

1  normal  LiCl 73 . 8 

0.5  normal  CaCl2 : 71.4 

0.5  normal  BaCl2 72.2 

0.5  normal  SrCla 72 

Some  of  the  magnesium  salts  may  completely  inhibit  heat  coagu- 
lation; MgCl2  below  6  normal,  Mg(N03)2  below  4  normal. 

Cations  also  may  be  divided  into  different  groups,  according  to 
their  influence: 

In  the  case  of  SO4,  CI,  Br  and  NO3,  there  is  a  greater  rise  with 
lower  concentrations  (up  to  0.5  to  about  1  normal)  then  a  smaller 
rise  up  to  1  normal.  In  the  case  of  SCN  and  I  the  inhibition  from 
1  to  2  normal  is  so  complete  that  no  coagulation  occurs  even  at  the 
highest  concentration  of  the  salt.  In  the  case  of  citrate,  acetate 
and  oxalate  the  coagulation  temperature  rises  sharply  from  0.05  to 
0.1  normal,  whereupon  the  curve  again  falls.  Obviously  this  is  as- 
sociated with  the  strong  hydrolytic  cleavage  of  these  weak  acids  in 


PROTEINS  151 

the  presence  of  strong  alkalis,  whereby  there  is  formed  more  or  less 
alkali  albumin  which  is  not  coagulated  by  heat. 

We  have  discussed  these  questions  separately  in  order  that  we  may 
obtain  a  picture  of  the  complicated  relations  which  also  reappear  in 
the  other  properties  of  albumin. 

Heat  coagulation  involves  two  overlapping  processes;  albumin  be- 
comes insoluble  and  it  flocculates.  Wolfgang  Pauli  and  H.  Hand- 
ovsKY  demonstrated  this  very  simply:  a  mixture  of  albumin  with 
2  normal  KSCN  was  boiled  and  a  portion  of  it  dialyzed  against  run- 
ning water.  The  control  portion  remained  clear,  but  the  portion  from 
which  the  KSCN  was  removed  by  dialysis  showed  marked  flocculation. 

A  further  influence  exerted  by  neutral  salt  upon  amphoteric  al- 
bumin is  the  change  in  viscosity,  the  internal  friction.  Although 
NaCl,  NaSCN,  Na2S04,  CaClz  and  KSCN  in  concentrations  of  0.01 
to  0.05  normal  raise  the  viscosity  of  water,  they  depress  that  of 
amphoteric  albumin  solution.  If  the  salt  concentration  rises,  the 
diminution  in  the  viscosity  of  albumin  may  finally  be  exceeded  by 
the  increase  in  the  viscosity  of  the  water,  as  occurs  in  fact  at  0.1 
normal  NaCl  and  (NH4)2S04.  Closer  observation  reveals  a  far- 
reaching  parallelism  between  the  influence  of  neutral  salts  on  heat 
coagulation  and  viscosity. 

If  non-neutral  salts,  or  salts  strongly  dissociated  hydrolytically, 
(Na3P04,  NaHCOs,  AICI3)  are  allowed  to  act  on  amphoteric  al- 
bumin, the  result  is  quite  different,  since  even  minute  traces  of  acid 
or  alkali  form  acid  or  alkali  albumins,  which  behave  quite  differently, 
as  we  shall  see.  With  higher  salt  concentration  the  albumin  is  salted 
out  or  flocculated.  Neutral  salts  of  the  alkalis  as  well  as  inagnesiujji 
cause  a  reversible  salting  out  such  as  occurs  also  with  salts  of  the 
alkaline  earths,  though  after  a  very  short  time  an  irreversible  coagu- 
lation sets  in.  Some  of  the  salts  of  the  heavy  metals  cause  an  im- 
mediate irreversible  coagulation.  With  the  alkali  salts,  the  cations 
(Li,  K,  Na,  NH4)  do  not  materially  differ  in  their  salting  out  action, 
but  the  anions  do,  as  may  be  seen  from  the  following  table  of  F. 
Hofmeister.  The  figures  refer  to  the  onset  of  turbidity  in  egg 
albumen  containing  globulin  but  according  to  Lewith  apply  also  to 

ox  serum:  MoIs  per  liter 

at  30-40°  C. 

Sodium  citrate 0 .  56 

Sodium  tartrate 0.78 

Sodium  sulphate 0 .  80 

Sodium  acetate 1 .  69 

Sodium  Chlorid 3. 62 

Sodium  nitrate 5 .  42 

Sodium  chlorate 5 .  52 

lodid  and  sulphocyanate  do  not  cause  precipitation. 


152  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Acid  Albumin. 

There  is  a  marked  change  in  the  properties  of  amphoteric  albu- 
min when  acid  is  added  to  it.  It  migrates  to  the  cathode  as  though 
it  were  the  basic  portion  of  a  salt;  it  loses  its  coagulability  by  heat 
and  alcohol,  its  internal  friction  is  greatly  increased  and  its  surface 
tension  diminished.  If  an  excess  of  acid  is  added,  coagulability  by 
acids  and  alcohol  is  restored  and  its  viscosity  diminishes. 

Sjöquist*  was  of  the  opinion  that  albumin  formed  with  acids 
strongly  hydrated  (swollen)  ionized  salts.  This  assumption  was  con- 
firmed by  the  researches  of  St.  Bugarszky  and  L.  Liebermann*  and 
of  K.  Spiro  and  Pemsel.*  It  was  finally  established  by  Mauabe 
and  J.  Matuta  by  extremely  accurate  measurements  on  the  ioniza- 
tion constants  of  acid  albumin.  W.  Pauli  and  M.  Hirschfeld  then 
established  that  albumin  was  polybasic,  i.e.,  behaved  like  a  tri-  or 
tetra-amino  acid,  and  that  the  salts  were  subject  to  the  normal  hydro- 
lytic  dissociation,  characteristic  of  weak  bases.  S.  Oden  and  W. 
Pauli  conclude  from  the  rise  in  migration  velocity  with  increasing 
fixation  of  acid  that  polyvalent  protein  ions  are  formed.^ 

In  a  solution  containing  about  1  per  cent  albumin,  the  maximum 
internal  friction  is  reached  at  0.016  normal  HCl,  and  falls  with 
greater  concentrations  of  acid.  Such  a  maximum  is  also  found  with 
other  acids  (oxalic  acid,  sulphuric  acid),  while  with  others  (acetic 
acid,  citric  acid)  a  continual  rise  in  internal  friction  accompanies  the 
concentration  of  the  acid. 

Precipitahility  hy  alcohol  runs  parallel  with  the  increase  or  decrease 
in  the  internal  friction  (K.  Schorr). 

When  amphoteric  albumin  has  been  made  incoagulable  by  acids, 
the  addition  of  neutral  salts  restores  the  coagulability  by  heat  and 
alcohol.  All  the  salts  investigated  (NaS04,  NaNOs,  Na3P04,  Na- 
acetate,  Na-formate,  etc.)  depress  the  internal  friction.  In  this  re- 
spect, the  cations  are  of  lesser  importance,  the  anions  being  decisive 
in  the  following  order: 

CI  <  NO3  <  SCN  <  SO4  <  C2H3O2. 

Nonelectrolytes  (cane  sugar,  urea)  have,  on  the  contrary,  little 
influence  in  this  respect. 

Caffein  and  its  salts  are  an  exception,  as  they  increase  the  internal 
friction  of  acid  albumin  (H.  Handovsky*^), 

An  excess  of  acid  alone  or  the  addition  of  neutral  salt  to  an  amount 
of  acid  which  is  insufficient  to  cause  precipitation  causes  at  first  a 

1  See  also  W.  E.  Ringer,  Acid  Fixation  by  Albumin  and  Viscosity,  Van 
Bemmelen-Festschrift  (Helder  i.H.u.  Dresden,  1910),  243-60. 


PROTEINS  153 

reversible  flocculation  of  albumin  in  the  cold,  but  with  greater  con- 
centration (from  about  0.03  normal  up)  an  irreversible  flocculation. 
Here  also  the  anions  have  an  unequal  influence,  which  is  arranged  in 
an  order  the  reverse  of  that  obtaining  for  neutral  albumin,  namely, 

SO4  <  CI  <  NO3  <  Br  <  SON. 

This  series,  accordingly,  does  not  agree  with  the  other  one  in  all 
respects. 

It  is  quite  evident  in  the  case  of  the  acid  salts  that  their  action  is 
the  combined  result  of  the  acid  albumin  formed  and  the  action  of 
the  salt  itself.     The  process  is,  therefore,  quite  complicated. 

Alkali  Albumin. 

There  is  a  far-reaching  parallelism  between  alkali  albumin  and 
acid  albumin.  Alkali  albumin  like  acid  albumin  is  not  coagulable 
by  heat  or  alcohol  (even  0.003  normal  NaOH  inhibits  the  heat  coagu- 
lation of  amphoteric  albumin) ;  its  viscosity  is  greatly  increased,  its 
surface  tension  diminished;  excess  of  alkali  restores  the  precipitability 
by  alcohol  and  again  decreases  the  internal  friction;  it  migrates  to  the 
anode.  St.  Bugarszky  and  L.  Liebermann*  showed  that  NaOH  was 
bound  by  albumin,  and  that  albumin  depressed  the  freezing  point  of 
soda-lye.  Neutral  salts  arrest  the  action  of  alkalis;  in  contradis- 
tinction to  acid  albumin  it  is  the  cations  to  which  the  greatest  signifi- 
cance attaches,  and,  in  fact,  the  effect  of  the  divalent  earth  alkalis 
(Ca,  Sr  and  Ba)  and  the  divalent  magnesimn  very  greatly  exceeds 
that  of  the  monovalent  alkalis.  Though  heat  coagulation  does  not 
occur  at  all  or  advances  only  to  a  milky  turbidity  (e.g.,  the  effect  of 
1.2  normal  KCl  was  doubtful),  in  alkali  albumin  containing  large 
quantities  of  alkaline  salts  the  ability  of  alkali  albumin  to  coagulate 
with  0.003  normal  NaOH  is  demonstrable  upon  the  addition  of 
0.0002  normal  CaClg. 

Additions  of  neutral  salts  bring  about  a  decrease  of  internal  friction 
in  a  manner  analogous  to  their  influence  on  heat  coagulation,  and,  in 
fact,  a  small  addition  of  salt  has  a  proportionately  greater  effect 
than  a  large  one.  Moreover,  the  earth  alkalis  greatly  exceed  the 
alkali  salts  in  their  ability  to  diminish  internal  friction. 

The  salting  out  of  alkali  albumin  requires  a  greater  concentration 
of  alkali  salts  than  is  required  for  neutral  albumin;  the  product  is 
reversible  and  the  anions  are  effective  in  the  same  order  as  for  neutral 
albumin. 

In  general,  the  relations  are  simpler  for  alkali  albumin  than  for 
acid  albumin.     In  the  former,  they  depend  upon  the  electrolytic  dis- 


154  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

sociation  of  the  base,  while  in  the  latter,  certain  electrochemical 
factors  which  may  not  be  disregarded  play  a  part. 

If  dilute  soda-lye  (0.025  normal  NaOH)  acts  for  a  long  time  on 
serum  albumin,  the  internal  friction  reaches  a  maximum,  remains 
constant  for  a  while  and  then  diminishes  (K.  Schorr).  Evidently 
there  occurs  fixation  of  water,  swelling.  The  cleavage  of  the  albu- 
min molecule  is  accompanied  by  the  formation  of  less  colloidal 
disintegration  products,  and  is  characterized  by  a  diminution  of  the 
viscosity. 

If  from  these  results  we  try  to  obtain  an  idea  of  the  processes  in- 
volved, we  shall  find  a  useful  guide  in  the  theory  of  the  amphoteric 
nature  of  genuine  albumin  proposed  by  G.  Bredig*  and  extended  by 
Wo.  Pauli.  Let  us  think  of  albumin  as  built  according  to  the 
structure  of  a  cyclic  ammonium  salt : 

R 
^COO 

in  which  R  represents  a  complicated  organic  complex  and  the  ab- 
sorption of  water  follows  according  to  the  scheme: 

yNHs  .NH3OH 

R  +  H.2O  <=±  R 

^COO  ^COOH 

This  is  an  amphoteric  electrolyte  which  unites  with  bases  and  acid, 
which  splits  off  H  as  well  as  OH  ions  and  in  which  the 

Ka  (acid  dissociation)  >  Kb  (base  dissociation) 

in  other  words,  it  behaves  like  a  very  weak  acid.     Pure  albumin 

consists  principally  of  electrically  neutral  particles  but  forms  acid 

and  alkali  salts  which  are  strongly  ionized. 

There  exist 

xNHaOH  ^NHsCl  ^NHsOH 

R  R  R 

^COOH  ^COOH  ^COONa 

neutral  albumin  acid  albumin  alkali  albumin 

That  the  albumin  ions  are  responsible  for  the  great  internal  friction 
is  to  be  assumed  from  the  investigations  of  E.  Laqueur  and  0. 
Sackur*  on  alkali-caseinates.  The  cause  of  this  phenomenon  is 
found  in  the  strong  hydration  (water  fixation,  swelling)  of  the  albu- 
min ions.  According  to  Wo.  Pauli  and  M.  Samec  the  existence  of 
polyvalent  ions  must  be  assumed  in  the  case  of  acid  and  alkali  albu- 


PROTEINS  155 

min.  Even  assuming  the  smallest  values  for  the  molecular  weight 
of  albumin,  the  quantities  of  acid  or  alkaU  found  are  so  large  that  they 
indicate  the  fixation  of  several  acid  or  alkali  molecules.  This  offers 
a  further  explanation  of  the  marked  increase  in  hydration  produced 
by  acids  and  alkalis.  The  stability  of  an  albumin  solution  and  its 
precipitability,  e.g.,  by  alcohol,  are  directly  proportional  to  the  num- 
ber of  albumin  ions  it  contains.  The  circumstances  here  are  quite 
analogous  to  those  with  crystalloids.  Ions  tend  to  go  into  solution 
and  to  form  hydrates;  the  saturation  concentration  of  neutral  par- 
ticles is  always  less  than  that  of  ions. 

In  this  way,  we  may  explain  the  properties  of  strongly  ionized 
pure  acid  and  alkali  albumin  as  contrasted  with  the  slightly  disso- 
ciated neutral  albumin.  How  does  this  theory  agree  with  the  effect 
of  neutral  salts?     "VYo.  Pauli  explains  it  in  the  following  way: 

,NH4C1  yNHaCl 

R  +  NaNOa  i^  R  +  HNO3 

^COOH  ^COONa 

Acid  albumin  +  neutral  salt       of  acid^lbllmin     +   ^^ee  acid 

In  this  way  was  explained  not  only  the  increased  number  of  free 
H  ions,  which  he  demonstrated,  but  also  the  marked  diminution  in 
internal  friction;  because  an  amphoteric  salt,  in  which  both  anions 
and  cations  tend  to  ionize  about  equally,  is  but  slightly  dissociated. 
The  action  of  neutral  salts  in  alkali  albumin  is  different;  it  follows 
the  following  scheme: 

yNHaOH  .NH2KCI 

R  +    KCl     ?=±  R  +H2O 

^COONa  ^COONa 

Alkali  albumin  +    neutral  salt    complex^albumin  ^  ^^^^^ 

Accordingly,  a  complex  albumin  salt  was  formed  to  which  a  less 
amount  of  ionization  may  be  ascribed  than  to  alkali  albumin.  The 
action  of  salts  of  the  alkaline  earths  follows  this  scheme: 

.NH3OH  .NHaNaNOs 

R  +  ^  NO3  :^  R  +  H2O 

^COONa  ^COO^ 

The  replacement  of  the  alkali  ion  in  the  carboxyl  of  the  amino 
group  results  in  a  weakly  ionized  complex  salt.  The  effect  on  albu- 
min of  organic  bases,  which  are  often  highly  toxic,  and  of  amphoteric 
electrolytes,  have  also  been  studied  by  H.  Handovsky,  and  the  re- 
sults agree  with  the  above  scheme. 


156  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  conditions  governing  the  action  of  neutral  salts  upon  acid 
albumin  are  not  sufficiently  understood  to  warrant  proposing  a  simple 
scheme.^ 

The  optical  rotation  of  albumin  runs  parallel  with  the  changes 
in  its  internal  friction  and  coagulability  (Wo.  Pauli,*^  M.  Samec, 
E.  Strauss).  In  fact,  the  albumin  ions  rotate  light  more  power- 
fully than  neutral  albumin. 

Let  us  summarize  briefly :  neutral  albumin  has  a  low  internal  fric- 
tion, coagulates  easily  and  shows  little  optical  rotation;  ionized  albumin 
has  high  internal  friction,  coagulates  with  difficulty  and  rotates  light 
powerfully;  neutral  salts  diminish  ionization. 

This  chemical  point  of  view  is  additionally  supported  by  the 
investigations  of  P.  Pfeiffer  and  J.  W.  Modelski  as  well  as  of  P. 
Pfeiffer  and  Wittka.  These  authors  have  shown  that  amino  acids 
and  Polypeptids  of  known  chemical  structure  form  with  neutral 
salts  of  the  alkalis  and  earth  alkalis,  crystalline  addition  compounds 
constructed  on  simple  stoichiometric  principles.  Some  of  these 
molecular  compounds  are  much  more  readily  soluble  in  water  than 
the  aminoacids  or  Polypeptids  and  some  much  less  soluble,  so  that, 
as  in  the  case  of  albuminous  substances,  it  is  possible  to  salt  some  of 
them  out  (analogous  to  globulins). 

Albumin  and  Inorganic  Hydrosols 

According  to  U.  Friedemann*^  electrolyte-free  albumin  is  precipi- 
tated both  by  positive  and  by  negative  inorganic  hydrosols.  Hydro- 
phobe hydrosols  such  as  AS2S3,  Au,  etc.,  regularly  form  precipitates, 
which,  according  to  W.  Pauli  and  Hecker,  are  not  inhibited  by  an 
excess  either  of  hydrosol  or  of  albumin.  Neutral  salts,  acids  and 
alkalis  exert  a  protective  action,  but  nonelectrolytes,  such  as  urea 
and  sugar,  are  inactive. 

In  the  case  of  positive  hydrophile  inorganic  hydrosols  such  as 
Fe  (OH)  3  there  is  an  optimum  precipitation  zone  that  lies  somewhere 
between  one  part  by  weight  of  Fe  (OH)  3  and  three  parts  by  weight 
of  the  electrolyte-free  albumin.  With  an  excess  of  Fe(0H)3  there  is 
increasing  solution  which  is  complete  in  about  the  proportion  of  two 
to  three;  there  is  no  complete  solution  with  an  excess  of  albumin. 
Neutral  salt  exerts  a  protective  action  when  albumin  is  in  excess  but 
on  the  contrary  favors  precipitation  when  Fe  (OH)  3  is  in  excess. 
Acids  inhibit  precipitation;   alkalis  precipitate  when  Fe  (OH)  3  is  in 

^  From  the  formula  it  should  not  be  assumed  that  only  free  terminal  NH2 
groups  are  considered.  As  the  result  of  the  work  of  Blasel  and  J.  Matuta  on 
deaminized  glutin  (glutin  whose  free  NH2  groups  are  satisfied)  it  is  more  probable 
that  its  interior  NH  groups  are  involved  in  the  formation  of  salts  with  acids. 


PROTEINS  157 

..•■ 

excess,  otherwise  they  exert  a  protective  action.  Hydrophile  negative 
inorganic  hydrosols,  e.g.,  siHcic  acid,  differ  from  positive  hydrosols 
only  by  the  presence  of  H  and  OH  ions  which  act  oppositely  to  those 
in  the  positive  hydrosols. 

Only  a  small  fraction  of  the  albumin  is  precipitated  by  hydrophobe 
inorganic  colloids;  but  the  greater  portion,  and  at  times  all  the  albu- 
min, is  precipitated  by  hydrophile  hydrosols. 

Albumins  appear  to  react  with  proteins  of  pronounced  basic  (histone) 
or  acid  character  (U.  Friedemann  and  H.  Friedenthal*)  just  as  do 
inorganic  hydrophile  hydrosols. 


Albumin,  Heavy  Metals  and  Salts  of  Heavy  Metals. 

On  shaking  salt-free  albumin  solutions  with  metallic  iron,  cobalt, 
copper,  lead,  nickel  or  aluminium,  portions  of  these  metals  go  into 
solution  and  are  bound  by  albumin  in  a  hitherto  unrecognized 
"masked"  form,  according  to  Benedicenti  and  Revello-Alves. 

Electrolyte-free  albumin  yields  no  precipitate  with  zinc,  copper, 
mercury  or  lead  salts.  In  the  presence  of  salts,  however,  albumin 
forms  with  salts  of  the  heavy  metals  precipitates  whose  chemical  com- 
position is  not  constant,  but  depends  on  the  concentration  of  the 
components  at  the  time  of  precipitation.  By  precipitating  albumin 
with  solutions  of  heavy  metal  salts  of  varying  concentrations  we 
get  ''irregular  series,"  which  frequently  show  two  zones  of  precipi- 
tation: one  with  very  dilute  solutions  of  the  metal  salt  (one  ten 
thousandth  normal  and  under)  and  another  with  high  concentration; 
between  these  there  is  always  a  zone  with  no  precipitation.  The 
precipitation  zone  with  great  dilutions  of  the  metal  salt  is  due  accord- 
ing to  H.  Bechhold*  to  metal  hydroxid  split  off  hydrolytically, 
which  precipitates  with  albumin,  forming  an  insoluble  heavy  metal- 
albumin  compound.  The  resolution  of  this  precipitate  at  somewhat 
greater  concentration  of  metal  salt  results  from  ionization.  W. 
Pauli  and  Hecker  have  shown  by  very  convincing  experiments 
upon  the  action  of  FeCls  on  albumin  that  a  soluble  ferric  ion-albumin 
complex  occurs  somewhat  in  accordance  with  the  following  scheme 
[a;Fe(OH)  3 -protein]  +  gFeCls  =  [xFe(OH)  3- protein]  gf  Fe  +  3gCl. 

Upon  addition  of  more  FeCls,  just  as  when  acid  is  added  to  acid 
albumin,  partial  neutralization  occurs  and  there  is  further  precipi- 
tation. Ur02Cl2  behaves  hke  FeCls,  as  do  also,  to  a  certain  extent, 
AgN03,  ZnS04  and  Pb(N03)2.  On  the  other  hand,  the  precipitate 
disappears  with  higher  concentration  of  CuCi2  and  HgCl2,  and,  abso- 
lutely no  precipitate  is  formed  with  electrolyte-free  albumin  and  the 
chlorides  of  Fe",  Co",  Mn",  Cd". 


158  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Globulin. 

Those  proteins  which  are  insoluble  in  pure  water  and  soluble  in 
salt  solutions  are  called  globulins.  They  are  constituents  of  the 
blood  serum^  eggs  and  milk  of  animals.  They  occur  in  other  organs 
in  traces,  thus,  e.g.,  thyreo-globulin,  the  iodin-containing  protein  of 
the  thyroid,  is  a  globulin.  Large  quantities  of  globulin  are  found 
stored  in  the  seeds  of  plants.  A  seed  globulin,  edestin,  has  been 
obtained  in  crystalline  form. 

If  serum  is  dialyzed  against  pure  water,  globulin  will  be  precipitated 
as  the  content  of  the  dialyzer  cell  (globulin)  parts  with  salt.  By 
ultrafiltration,  H.  Bechhold*^  was  able  to  separate  globulin  from  the 
common  salt  holding  it  in  solution.  Globulins  are  also  soluble  in  acids 
and  alkalis.  If  globulins  are  kept  undissolved  (e.g.,  dried  or  sus- 
pended in  distilled  water)  a  change  occurs ;  they  lose  more  and  more  of 
their  solubility  in  dilute  solutions  of  neutral  salts.  Like  the  albumins, 
globulins  are  amphoteric:  without  the  presence  of  salt  they  have  no 
definite  direction  of  migration;  in  the  presence  of  traces  of  alkali 
they  pass  to  the  anode  and  in  the  presence  of  acids  they  pass  to  the 
cathode.  According  to  L.  Michaelis,  an  H  ion  concentration  of 
4.10"'^  is  the  isoelectric  point  for  serum  albumin.  According  to  W.  B. 
Hardy,  *^  a  given  quantity  of  salt-free  globulin  is  dissolved  by  an 
equimolecular  quantity  of  strong  monobasic  acids  (HCl,  HNO3, 
monochloracetic  acid).  The  weaker  the  acid  the  more  of  it  is  neces- 
sary to  dissolve  the  globulin.  About  twice  as  much  sulphuric  acid, 
tartaric  acid  and  oxahc  acid,  and  three  times  as  much  phosphoric 
acid  and  citric  acid,  is  required  than  of  HCl.  W.  B.  Hardy  concludes 
from  this  that  globulins  form  salts  with  acids  which  in  the  case  of 
weak  acids  are  greatly  hydrolyzed. 

Bases  act  in  a  manner  similar  to  the  acids,  with  the  exception 
that  NH3  dissolves  as  much  globulin  as  NaOH. 

Rise  of  temperature  increases  the  hydrolysis,  i.e.,  globulin,  dis- 
solved in  an  amount  of  weak  acid  or  weak  alkali  just  sufficient  to 
give  a  clear  solution,  becomes  turbid  when  it  is  warmed;  however, 
the  process  is  not  completely  reversible. 

It  was  deduced  from  the  conductivity  values  of  alkali  globulin  that 
globulin  is  a  pentavalent  acid,  and  from  its  saponification  with 
methyl  acetate  as  well  as  its  action  in  the  inversion  of  cane  sugar, 
that  it  is  of  a  more  strongly  acid  than  basic  character.  This  is  also 
evident  from  the  fact  that  the  conductivity  of  its  acid  salts  increases 
progressively  more,  when  diluted,  than  the  conductivity  of  its  alkali 
salts.  The  preponderant  acid  character  is  also  evident  from  the  fact 
that  litmus  is  reddened  by  globulin. 


PROTEINS  159 

.J 

As  in  the  case  of  albumin,  the  globulin  ions  are  responsible  for  the 
internal  friction.  Though  the  internal  friction  of  globulin  in  neutral 
salts  is  low,  it  is  considerably  higher  in  the  ionized  solutions  occurring 
in  acids  or  alkalis;  the  viscosity  is  highest  in  the  case  of  alkali 
salts  of  globulins  which  are  ionized  most  strongly  and  least  hydrolyzed. 
The  viscosity  rises  disproportionately  with  concentration  and,  in  fact, 
the  increase  for  alkali  globulin  >  for  acid  globulin  >  for  neutral  salt- 
globulin  (W.  B.  Hardy*2). 

W.  B.  Hardy  gives  the  following  viscosity  values  for  7.59  gm. 
globulin  per  liter: 

Water 1 

MgS04-globulm 4.66 

HCl-globuUn 15.5 

NaOH-globuUn 67.9 

He  derived  these  velocities  for  globulin  ions: 

Acetic  acid-globulin 23  •  10~^  cm.  per  second 

HCl-globuUn 10-10^    "       " 

NaOH-globuUn 7.7-10-^    "      "        " 

W.  B.  Hardy  regards  solutions  of  globulin  in  neutral  salts  as 
molecular  combinations,  since,  in  contrast  to  solutions  in  alkalis  or 
acids,  they  are  thrown  down  upon  dilution.  It  can  be  understood 
from  the  dominant  acid  character  of  globulins  that  a  neutral  salt 
solution  of  globulins  is  precipitated  by  acids.  Though  alkali  globulin 
solutions  are  permanent  in  the  presence  of  neutral  salts,  acid  globu- 
lins are  precipitated  by  them. 

According  to  W.  B.  Hardy,  serum  contains  no  globulin  ions. 

If  serum  is  kept  warm  for  a  long  time  {e.g.,  2  hours)  below  its 
coagulation  temperature,  the  amount  of  globulin  is  increased  at  the 
expense  of  the  albuminous  portion  (Moll*).  This  formation  of 
globulin  is  either  impeded  or  entirely  stopped  by  salts. 

"Artificial  globulins"  is  the  designation  of  the  substances  pre- 
pared from  egg  albumin  by  Andre  Mayer.*  He  found  that  when 
he  added  to  egg  albumen  a  certain  quantity  of  a  solution  of  a  salt  of 
a  heavy  metal  ZnS04,  Zn(N03)2  or  a  positive  colloid  (colloidal  Fe203), 
the  resulting  precipitate  was  insoluble  in  water  and  in  solutions  of 
nonelectrolytes,  but,  on  the  contrary,  it  was  soluble  m  solutions  of 
salts  (e.g.,  NaCl,  Ca(N03)2,  etc.).  With  these  tacts  in  mind,  we  must 
consider  the  suggestion  made  by  A.  Mayer  that  globulins  are  com- 
plexes of  albumins  (possibly  with  other  positive  colloids). 


160  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Fibrin. 

Fibrin  is  the  substance  of  blood  plasma,  which  coagulates  shortly 
after  the  blood  has  left  the  vessels.  Upon  the  clotting  of  plasma, 
which  contains  no  blood  corpuscles,  no  jelly  is  formed,  but  character- 
istic fibrous  masses.  Formerly  it  was  thought  that  uncoagulated 
fibrin,  called  fibrinogen  (see  p.  299),  was  something  quite  different 
from  fibrin.  As  a  result  of  the  investigations  of  Hekma  it  is  possible 
that  fibrinogen  is  the  hydrosol  of  alkali  fibrin.  If  fibrin  is  dissolved 
in  extremely  dilute  alkali  we  obtain  a  fluid  having  all  the  properties 
of  fibrinogen.  Normal  coagulation  outside  the  blood  vessels  as  well 
as  the  resulting  product  must  be  sharply  differentiated  from  fibrin 
coagulated  by  heat.  Fibrin  coagulated  by  heat  ceases  to  show  the 
swelling  phenomena  it  possessed  before  it  was  heated;  it  has  become 
hydrophobe.  When  coagulated,  fibrin  is  an  irreversible  gel.  In 
weak  acid  and  alkalis  it  swells  and  gradually  goes  into  solution 
following,  as  it  does  so,  the  same  laws  as  does  gelatin  (see  p.  68, 
et  seq.).  Martin  H.  Fischer*  has  studied  its  swelling  under  the  in- 
fluence of  acids,  bases  and  salts,  and  utilized  his  results  for  his  theory 
of  edema  (see  p.  223,  et  seq.) 

Muscle  albumin  or  myosin,  the  coagulation  of  which  at  death 
causes  rigor  mortis,  belongs  to  the  same  group  as  fibrin. 

Nucleins. 

Basic  substances  have  been  prepared  from  cell  nuclei;  histone 
from  the  leucocytes  of  the  thymus,  fish  roes,  etc.,  as  well  as  pro- 
tamine, so  thoroughly  studied  by  A.  Kossel  and  usually  obtained 
from  the  spermatozoa  of  several  different  kinds  of  fish.  They  do 
not  exist  as  such  in  these  organs  but  occur  in  combination  with  acid 
nucleins  as  nucleo-proteins  and  nucleo-histones. 

Neutral  solutions  of  histone  yield  a  precipitate  containing  very 
little  salt  with  solutions  of  egg  albumin,  casein  and  serum  globulin. 
When  we  recall  that  casein  and  globulin  are  of  decided  acid  reaction, 
their  union  with  basic  histone  is  quite  easily  understood.  A  priori, 
it  is  improbable  that  the  precipitate  should  contain  1  part  histone, 
2  parts  casein  and  globulin  and  1  part  egg  albumin,  as  has  been 
claimed.  It  has  been  shown  by  U.  Friedemann  and  H.  Frieden- 
thal* that  according  to  the  relative  concentration  in  which  solutions 
of  histone  and  albumin  are  mixed,  the  precipitate  will  vary  in  com- 
position; that  the  addition  of  NaCl  changes  the  precipitation  limits 
and  that  fresh  solutions  have  different  precipitation  limits  than 
older  ones.  All  these  facts  point  with  certainty  to  the  fact  that 
nuclein  is  not  a  definite  chemical  combination,  but  that  nucleins  are 
colloid  compounds  consisting  of  a  negative  and  a  positive  colloid. 


PROTEINS  161 

Albuminoids.     (Scleroproteins) 

Though  the  organic  framework  of  plants  consists  of  cellulose, 
that  of  animals  is  formed  of  nitrogenous  substances  classified  as 
albuminoids.  Like  cellulose,  they  are  very  resistant  chemically  to 
foreign  influences,  water,  salt  solutions,  acids  and  bases. 

The  most  important  of  the  albuminoids  is  collagen,  derived  from 
bone,  cartilage  and  the  fibrils  of  connective  tissue.  On  boiling  with 
water,  it  swells  and  gradually  dissolves,  undergoing  hydrolytic  cleav- 
age and  forming  glue  or  gelatin.  Gelatin,  which  has  been  the  subject 
of  the  most  important  investigations  concerning  hydrophile  gels  and 
from  which  the  whole  class  of  gels  take  their  name,  does  not  occur  in 
the  organism  at  all.  The  most  important  data  concerning  it  have 
been  given  on  page  68,  et  seq.  What  has  been  said,  especially  in 
reference  to  the  preparation  of  a  solution  of  agar  (p.  137)  holds  for 
gelatin  as  well.  It  should  be  recalled  that  acids  and  alkalis  greatly 
increase  the  swelling  of  gelatin.  The  swelling  capacity  reaches  a 
maximum  with  increasing  concentration  of  HCl  (0.025  n)  and  KOH 
(0.028  n)  (Wo.  Ostwald).  We  thus  find  an  absolute  parallelism 
between  the  swelling  of  gelatin  and  the  ionizati(?n  of  albumin  (see 
pp.  152  to  156).  In  excellent  agreement  with  this  is  the  fact  that 
the  minimal  swelling  occurs  at  the  isoelectric  point  of  gelatin,  namely, 
with  an  H  ion  concentration  of  2.10"^  (L.  Michaelis,  R.  Chiari).  It 
must  be  emphasized  especially,  that  a  very  dilute  solution  of  gela- 
tin depresses  (according  to  G.  Quincke)  the  surface  tension  of  water 
12  per  cent.  The  solubiHty  of  CO2  is  very  considerably  greater  in 
gelatin  sols  than  in  water  (in  contrast  to  other  hydrophile  sols). 

Compared  with  other  colloids  (serum  albumin),  gelatin  lowers  the 
solubility  of  easily  soluble  electrolytes  and  increases  that  of  those 
soluble  with  difficulty.  The  following  are  the  figures  from  the  in- 
vestigations of  Wo.  Pauli  and  M.  Samec:* 

There  dissolves  in  100  gm    water  +  i  per  cent  gelatin  +  10  per  cent  gelatin 

Ammonium  chlorid 28.49  27.55  26.48 

Magnesium  chlorid 35.94  35.22  35. 13 

Ammonium  sulphocyanate 62.46  61.46  58.92 

1.5  per  cent  gelatin 

Calcium  sulphate 0. 223  0.295 

Tertiary  calcium  phosphate 

Ca3(P04)2 0.011  0.018 

Calcium  carbonate 0.004  0.015 

Silicic  acid 0.023  0.027 

The  solidification  and  the  melting  points  depend  greatly  upon  the 
previous  history  of  the  gelatin;  the  longer  gelatin  is  warmed  the 
less  it  tends  to  solidify.     Upon  heating  a  2  per  cent  gelatin  solution 


162  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

to  100°  C,  the  relative  internal  friction  (according  to  P.  von  Schroe- 
der)  falls  from  1.75  (at  the  end  of  one-half  hour)  to  1.22  (at  the  end 
of  16  hours).  Possibly  this  is  due  to  the  increasing  hydrolytic  cleav- 
age.    The  following  figures  give  some  idea  of  the  relations: 


Content  per  liter. 

Solidification  temperature,  °  C. 

Melting  temperature,  °  C. 

Grams. 
1.8 
2.5 
50 

100 

150 

<10      (Rohloff  and  Schinja) 
0      (S.  J.  Levites) 
17.8  (Pauli  and  Rona) 
21      (Pauli  and  Rona) 
25 . 5  (Pauli  and  Rona) 

26.1  (Pauli  and  Rona)* 
29.6  (Pauli  and  Rona) 
29.4  (Pauli  and  Rona) 

These  solidification  temperatures  are  markedly  shifted  by  elec- 
trolytes and,  in  fact,  the  anions  have  the  greatest  influence,  whereas 
the  cations  are  of  less  moment. 

The  solidification  temperature  is  raised  by  )  SO4  >  CH3CO2  > 

The  solidification  time  is  shortened  by        )     tartrates. 

^,        Ti-r.     .•      ^  .        •   ,  n     ibenzoates    and    salicy- 

The  solidification  temperature  IS  lowered  by      ,  ^     ^  o/^tvt^  t^ -n 
rru       vA-ti    V      +•       •    1       +V.      Au       \    lates>SCN>I>Br 
The  solidification  time  is  lengthened  by  .^  ^^^   ^    r^^ 

^  -^     ]     >  NO3  >  CI. 

The  following  data  (from  H.  Bechhold  and  J.  Ziegler*^)  serve 
as  an  example: 

Melting  point. 

10  per  cent  gelatin 31.6 

10  per  cent  gelatin  +  1  mol.  NaCl 28. 5 

10  per  cent  gelatin  +  2  mol.  Na2S04 34. 2 

10  per  cent  gelatin  +  1  mol.  Nal 10.0 

Nonelectrolytes  also  influence  the  melting  point  of  gelatin. 
Glycerin  and  sugar  (mannit,  cane  sugar,  etc.),  in  contradistinction 
to  agar,  raise  the  temperature  and  increase  the  rate  of  gelatinization, 
while  furfurol,  urea,  alcohols,  resorcin,  hydrochinon  and  pyrogallol 
lower  them.  Nongelatinizing  colloids  have  no  influence  on  gelatini- 
zation. 

The  following  figures  from  H.  Bechhold  and  J.  Ziegler*^  serve 
to  make  this  clear: 

Melting  point. 

10  per  cent  gelatin 31 .  66 

10  per  cent  gelatin  +  1  mol.  grape  sugar 32 .  25 

10  per  cent  gelatin  +  2  mol.  glycerin 32. 17 

10  per  cent  gelatin  +  2  mol.  alcohol 30 . 0 

10  per  cent  gelatin  +  1  mol.  urea 26. 3 

Precipitation  of  the  gelatin  sol  must  be  sharply  differentiated  from 
gelatinization.  Precipitation  is  induced  by  electrolytes,  whereas 
nonelectrolytes  usually  interfere  with  it.     Precipitation  corresponds 


PROTEINS  163 

rather  to  salting  out,  which,  we  may  assume,  occurs  also  in  the  case 
of  crystalloids;  it  may  be  induced  not  only  by  electrolytes  which 
raise  the  melting  point  of  the  gel,  but  also  by  those  which  depress  it. 
Precipitation  becomes  evident  at  first  through  a  turbidity  which 
may  be  sufficiently  marked  to  give  a  tenacious  gelatin  phase  and  a 
more  limpid  aqueous  phase.  In  precipitations,  also,  the  anions  have 
the  determining  influence,  their  precipitating  effect  being  arranged  in 
the  following  order: 

SO4  >  Citrate  >  Tartrate  >  Acetate  >  Chlorid. 

Inorganic  hydrosols  behave  quantitatively  toward  gelatin  the 
same  as  toward  albumin  (see  p.  156). 

The  swelling  and  shrinking  of  gelatin  referred  to  on  page  68,  et  seq., 
are  characteristic  for  all  elastic  gels. 

According  to  J.  Traube  and  F.  Köhler  there  exists  a  parallelism 
between  the  swelhng,  shrinking,  solidification  and  melting  point  of 
gelatin  when  it  is  mixed  with  other  substances. 

The  tinctorial  properties  of  elastic  fibers  and  their  chief  constituent, 
elastin,  are  better  known  than  their  other  colloidal  properties. 

To  the  investigations  of  P.  G.  Unna  and  L.  Golodetz,  we  owe  our 
knowledge  of  the  keratins,  the  horny  substances  composing  skin, 
hair,  nails,  hoofs,  horns,  feathers,  etc.  Chemical  studies  of  these 
substances  are  very  difficult  because  they  resist  chemical  attack. 

We  shall  merely  mention  the  remaining  albuminoids,  s-pongin,  the 
structural  support  of  ordinary  sponge,  chonchiolin,  the  framework  of 
mussels  and  snails  and  further,  the  albumoids,  a  group  into  which 
almost  all  unclassified  proteins  are  thrown. 

Nucleoalbumins. 

These  proteins,  like  the  albumins,  are  digested  by  pepsin-hydro- 
chloric acid;  they  dissolve  almost  entirely,  but  at  the  same  time 
split  off  an  almost  insoluble  phosphorus-containing  complex.  The 
casein  of  milk,  the  vitellin  of  egg  yolk  and  perhaps  also  legumin  and 
vegetable  casein  are  nucleoalbumins.  It  is  remarkable  that  among 
the  phosphorus-containing  proteins  obtained  from  seeds,  there  should 
be  several  that  are  soluble  in  alcohol  (gliadin  from  cereal  grains  and 
zein  from  corn).  Because  of  this,  it  is  a  question  however,  whether 
they  are  related  to  casein. 

On  account  of  its  importance,  casein  has  been  most  extensively 
investigated.  In  milk,  casein  exists  as  a  salt  (united  to  lime  and 
alkali)  and,  being  dissolved,  exhibits  profound  hydrolytic  dissociation. 
Casein  may  be  thrown  out  of  solution  by  the  addition  of  acids  or  ren- 


164  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

nin,  yet  the  casein  obtained  by  the  addition  of  acids  and  that  obtained 
by  rennin  are  not  identical.  Furthermore,  casein  may  be  separated 
from  the  crystalloid  portions  of  milk  by  ultrafiltration  (H.  Bechhold 
and  by  centrifugation  (H.  Friedenthal*).  We  are  indebted  to  E. 
Laqueur  and  0.  Sackur*  as  well  as  T.  B.  Robertson*  (who  gives  a 
bibliography)  for  the  exhaustive  chemical  studies  of  casein  upon  which 
we  base  our  remarks.  In  water,  casein  is  completely  insoluble  and 
decidedly  acid.  A  piece  of  casein  stains  damp  blue  litmus  paper  red. 
According  to  L.  Michaelis  the  isoelectric  point  occurs  when  the  H 
ion  concentration  is  2.10"^.  Casein  forms  salts  soluble  in  water  with 
alkahs  and  alkaline  earths.  One  grain  of  casein  binds  8.81  c.c.  1/10 
normal  alkali  (using  Phenolphthalein  as  an  indicator).  From  this 
the  combining  weight  of  casein  is  1135,  and  a  common  multiple  of 
this  is  its  molecular  weight.  E.  Laqueur  and  0.  Sackur  deduced 
from  the  conductivity  of  sodium-casein  solution  with  increasing 
dilution,  that  casein  was  a  tetra-  or  hexabasic  acid  and  that,  therefore, 
its  molecular  weight  lay  between  4540  and  6810.  T.  B.  Robertson, 
as  a  result  of  his  investigations,  comes  to  the  contrary  conclusion, 
that  only  a  single  carboxyl  group  is  available  for  union  with  a  base. 
W.  van  Dam*  has  investigated  the  diminution  in  H  ion  concentration 
of  lactic  acid  solution  upon  adding  casein  and  concludes  from  it,  that 
a  basic  group  unites  with  four  replaceable  H  atoms  in  a  casein  mole- 
cule. 

In  solution,  casein  salts  are  hydrolytically  dissociated  and,  in 
fact,  it  follows  from  the  following  experiment  that  casein  (acid) 
forms  a  hydrosol.  A  neutral  solution  of  casein-sodium  solution  is 
slightly  opalescent  and  becomes  clear  upon  the  addition  of  an  alkali. 
The  solution  of  casein-lime  salts  are  still  more  opalescent  since  the 
alkaline  earth  salts  are  weaker  bases.  Casein-sodium  does  not 
diffuse  through  parchment;  the  membrane  must  have  a  decided 
difference  in  potential  since  the  sodium  ion  has  a  strong  tendency  to 
diffuse. 

E.  Laqueur  and  0.  Sackur  showed  that  the  internal  friction  of 
casein  salt  solutions  increased  proportionately  to  the  electrolytic 
dissociation,  and  that  every  diminution  of  electrolytic  dissociation 
was  accompanied  by  a  diminution  of  internal  friction.  The  casein 
ions  are  thus  responsible  for  high  internal  friction. 

Hemoglobin. 

Hemoglobin,  the  coloring  matter  of  blood,  has  only  recently  been 
studied  by  P.  Bottazzi.  It  is  preeminently  suited  for  colloid-chemi- 
cal investigation  on  account  of  its  color,  ease  of  crystallization  and 


PROTEINS  165 

pronounced  colloid  character.  Chemically,  it  is  composed  of  the 
protein  glohin,  a  histone,  and  the  iron-containing  component,  hematin, 
which  is  apparently  a  pyrrol  derivative. 

Bechhold  used  1  per  cent  hemoglobin  solutions  to  gauge  his 
ultrafilters  (see  p.  99). 

After  dialyzing  three  to  four  months,  hemoglobin  solutions  have  a 
conductivity  of  K  20°  =  1  X  10~*.  After  dialyzing  five  and  a  half 
months,  the  hemoglobin  was  completely  precipitated  though  the  pre- 
cipitate did  not  have  the  amorphous  flocculent  character  of  other  pro- 
teins but  was  more  granular  although  no  crystalline  formations  could 
be  recognized.  If  the  granules  were  removed  by  filtration  during  the 
dialysis,  there  was  obtained  a  reddish,  optically  inactive  solution 
which  showed  no  particles  in  the  ultramicroscope.  Such  a  solution 
does  not  pass  through  the  dialyzing  membrane  and  contains  particles 
which  are  somewhat  larger  than  those  of  serum  albumin  as  deter- 
mined by  ultrafiltration. 

Regarding  the  absorption  of  O  and  CO2  by  hemoglobin  see  page 
308,  et  seq. 

During  dialysis,  hemoglobin  changes  to  methemoglobin.  Methe- 
moglobin,  which  is  insoluble  in  water  and  neutral  salts,  redissolves 
upon  the  addition  of  traces  of  alkalis  or  acids. 

Purified  hemoglobin  migrates  to  the  anode.  In  view  of  this  fact 
and  the  relatively  high  conductivity  of  a  dialyzed  hemoglobin  solution 
P.  BoTTAZzi  assumes  that  hemoglobin  is  an  hemoglohinic  acid  in- 
soluble in  water,  but  which  exists  in  solution  as  an  alkali  hemoglo- 
binate.  Being  an  amphoteric  electrolyte  it  is  also  soluble  in  acids  and 
then  migrates  to  the  cathode;  its  H  ion  dissociation  far  exceeds  its 
OH  ion  dissociation.  According  to  L.  Michaelis,  on  the  contrary, 
hemoglobin  is  less  acid  than  serum  albumin;  its  isoelectric  point 
occurs  with  an  H  ion  concentration  of  1,8.10"''.  The  viscosity  curves 
which  P.  BoTAZzi*^  obtained  on  dissolving  it  in  alkalis  and  acids 
indicate  the  occurrence  of  methemoglobin  ions;  they  resemble  the 
viscosity  curves  of  alkali  and  acid  albumin.  Completely  dialyzed 
methemoglobin  coagulates  at  47°-53°C.;  in  the  presence  of  traces 
of  alkalis  or  acids,  and  in  the  absence  of  neutral  salts  coagulation 
fails  to  occur  even  at  100°  C. 


In  conclusion  we  shall  mention  the  mucins  and  mucoids.  They 
are  the  excretory  products  of  many  glands  and  may  be  briefly  de- 
scribed as  animal  mucus.  The  possession  of  a  carbohydrate  in  ad- 
dition to  the  protein  component  distingufshes  them  chemically. 
CoUoid-chemically  they  also  occupy  an  intermediate  position,  since 
they  are  not  coagulated  by  heat  but  are  precipitated  by  salts  and 
alcohol.  Because  of  their  acid  character  they  are  precipitated  by 
acids  and  dissolved  by  alkalis. 


166 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


The  Colloid  Cleavage  Products  of  Proteins. 

Proteins  undergo  hydrolytic  cleavage  by  acids,  alkalis  and 
enzymes.  From  the  viscosity  curve  resulting  from  the  prolonged 
action  of  NaOH  on  albumin,  it  is  deduced  that  the  disintegration 
of  albumin  depends  upon  the  albumin  ions  (W.  M.  Bayliss,  K. 
Schorr).  T.  B.  Robertson  arrived  at  a  similar  conclusion  from  his 
studies  of  the  tryptic  digestion  of  casein.  Under  these  circum- 
stances, it  is  obvious  that,  in  general,  the  digestion  of  albumin  by 
enzymes  occurs  more  readily  in  acid  or  alkaline  solution  than  in 
neutral  solution  where  there  are  but  few  albumin  ions. 

With  increasing  subdivision  of  the  molecule,  the  diffusibility,  etc., 
increases,  and  the  precipitability  by  neutral  salts  decreases.  The 
group  of  cleavage  products,  known  as  alhumoses,  dialyze  slowly 
through  animal  membranes,  whereas  the  peptones  are  not  to  be  dis- 
tinguished in  this  respect  from  true  crystalloids.  That  they  are 
still  to  a  certain  extent  colloids  is  proved  by  their  forming  films  on 
the  surface  of  water  (described  on  p.  33)  which  places  them  in  the 
class  of  those  dyes  which  lie  midway  between  colloids  and  crystal- 
loids. H.  Bechhold  arrived  at  a  similar  result  by  separating  albu- 
moses  from  the  remaining  fluid  by  means  of  ultrafiltration,  whereas 
peptones  and  the  closely  related  deuteroalbumoses  C  were  not  held 
back  even  by  10  per  cent  filters. 

The  albumoses  and  perhaps  also  the  peptones  are  evidently  mix- 
tures of  numerous  different  substances  which  have  not  yet  been 
chemically  identified.  They  are  differentiated  and  classified  ac- 
cording to  their  precipitability  by  electrolytes  and  alcohol,  which 
doubtless  stands  in  a  certain  relationship  to  the  size  of  their  molecules 
and  particles  and  to  their  ionization,  see  page  152.  By  means  of  ultra- 
filters  of  different  permeability,  H.  Bechhold  separated  albumoses 
into  "various  groups  which  corresponded  to  their  precipitability. 

Subjoined  is  the  classification  of  F.  Hofmeister-Pick  of  the  results 
by  ultrafiltration: 


Hetero-  and  protoalbu- 

moses 

Deuteroalbumoses  A . . . 
Deuteroalbumoses  B . . . 

Deuteroalbumoses  C. . . 

Peptones 


Saturation  per  cent 
of  ammonium  sul- 
phate required  to 
produce  precipita- 
tion. 


J        24-42 

54-62 
70-95 

100  +  acid 

Not  salted  out 


Ultrafilter, 
2.5  per  cent. 


3 

10  (H4) 

10 


Saturation  per  cent  of  am- 
monium sulpliate  required 
to  produce  precipitation. 


Residue  23 

Filtrate  34 

Residue  "    34 

Filtrate  95-100 

Residue  95-100+  acid, 
filtrate,  small  frac- 
tion 

Pass  through  filter 


PROTEINS  167 

These  experiments  were  undertaken  and  largely  carried  out  by 
Edgard  Zunz.*^  They  lead  to  a  multitude  of  valuable  results. 
It  suffices  to  mention  that  by  ultrafiltration  of  thioalbumose/  there 
were  shown  to  be  two  components  of  obviously  different  chemical 
constitution;  and  that  in  hetero-  and  protoalbumose  at  least  two  and 
in  the  latter  probably  even  three  ''proteoses"  should  be  assumed. 

^  Deuteroalbumose  A,  on  account  of  its  high  content  of  easily  spht-off  sul- 
phur, is  termed  "  Thioalbumose  "  by  Pick. 


CHAPTER  XI. 
FOODS  AND   CONDIMENTS. 

Formerly  the  preparation  of  food  was  one  of  the  most  important 
tasks  assigned  to  the  housewife;  nowadays  among  the  middle  and 
better  classes  this  duty  is  almost  entirely  surrendered  to  servants, 
while  among  the  working  classes  the  women  can  give  it  but  little  at- 
tention as  they  must  increase  the  family  income  by  work  away  from 
home.  These  conditions  have  brought  with  them  the  steady  de- 
terioration of  the  Art  of  Cookery.  The  raw  materials  nowadays 
supplied  to  the  kitchen  from  wholesale  establishments,  e.g.,  the 
bread,  fruit,  vegetables,  beer,  and  perhaps  even  meat,  etc.,  are,  it  is 
true,  of  a  much  superior  quality  than  formerly.  This  is  due  to  com- 
petition, easier  means  of  communication,  improvement  in  methods 
of  cultivation  and  all  the  advantages  consequent  upon  production 
on  a  large  scale.  The  conversion  of  this  raw  material  into  palatable 
meals  requires  a  large  measure  of  experience,  loving  care,  and  great 
interest  —  which  one  can  expect  from  neither  a  twenty-year-old  cook 
nor  the  tired  working  woman. 

Nutrition  is  undoubtedly  the  most  important  factor  in  our  whole 
social  life;  if  we  place  the  yearly  expense  for  nourishment  in  the 
German  Empire  at  ten  milhards  of  marks,  it  is  surely  underestimated. 
Only  a  one  per  cent  increase  of  the  successful  utilization  of  food 
would  show  a  yearly  profit  of  at  least  one  hundred  million  marks 
($25,000,000). 

It  is  hardly  to  be  expected  that  we  shall  accompUsh  this  by  re- 
turning to  former  conditions,  but  rather,  in  a  different  way,  namely, 
the  development  of  the  art  into  the  science  of  cookery.  The  kitchen 
will  probably  adapt  itself  more  and  more  to  wholesale  prepara- 
tion, and  then  there  will  be  men  and  women  who  will  choose 
cooking  as  a  profession  because  of  their  scientific  training  for  the 
work. 

Colloid  Chemistry  furnishes  us  the  rules  for  the  selection  and 
preparation  of  our  foodstuffs;  because  cooking  is  nothing  but  prac- 
tical colloid  chemistry.  Our  foodstuffs  consist  entirely  of  colloids 
and  their  nutritive  value  is  to  be  judged  mainly  from  a  colloid-chemi- 
cal point  of  view. 

168 


FOODS  AND   CONDIMENTS  169 

Very  little  truly  scientific  work  ^  has  as  yet  been  accomplished  in 
this  field;  so  that  we  must  content  ourselves  with  indicating  the 
problem.* 

Meat.  What  we  consume  as  ''  meat  "  consists  for  the  most  part 
of  muscle  fibers  and  connective  tissue  with  the  interspersed  fat.  In 
judging  the  meat  of  healthy  animals  its  source  is  the  chief  criterion; 
young,  well-nourished  animals  possess  a  juicy  meat  and  tender  con- 
nective tissue,  whereas  old  worn-out  animals  are  less  juicy  and  their 
connective  tissue  shows  a  firmer  structure.  From  these  few  premises 
it  is  evident  that  it  is  a  question  of  turgor  and  swelling  capacity, 
which  change  with  age;  this  is  an  important  colloid-chemical  prob- 
lem. It  is  still  an  open  question  whether  the  toughening  of  con- 
nective tissue  may  be  compared  to  the  lignification  of  the  vascular 
bundles  of  plants,  which  according  to  H.  Wislicenus*  (see  pp.  249 
and  250)  is  due  to  the  adsorption  of  colloids  from  the  cambial  sap. 

Fresh  killed  meat  is  tender;  it  becomes  soft  again  only  upon  the 
disappearance  of  rigor  mortis.  This  is  conditioned  by  phenomena 
of  swelling  and  shrinking,  an  interesting  method  for  studying  meat 
adopted  by  0.  von  Fürth  and  E.  Leuk.  After  death  lactic  acid 
accumulates  in  the  muscular  tissue,  and  greatly  increases  the  swell- 
ing capacity  of  the  muscles  (see  p.  290).  If  such  a  muscle  is 
placed  in  a  dilute  salt  solution  it  swells  up  and  after  about  25 
hours  has  taken  up  a  maximum  amount  of  water  of  swelling.  Then 
shrinking  occurs  as  the  result  of  the  progressive  coagulation  of  the 
muscle  albumin.  The  curve  obtained  in  this  manner  has  a  quite 
characteristic  shape  depending  on  what  has  happened  to  the  meat. 
Fig,  A  shows  the  curve  of  swelling  of  horse  heart  three  or  four 
hours  after  slaughter;  Fig.  B,  after  it  has  been  kept  3  days  in  the  ice 

1  In  this  connection  Dr.  J.  G.  M.  Bullowa  mentions  favorably  "The  Chemistry 
of  Cookery,"  by  Mattietj  Williams,  London,  Chatto  &  Windus,  1892.  [H.  C. 
Sherman  has  included  in  his  book,  "Food  Products,"  Macmillan  Co.,  1917,  the 
important  recent  data.  By  reason  of  the  war,  the  preservation  of  perishable  food- 
stuffs has  assumed  great  importance.  Dehydrating  processes  make  possible  the 
transportation  of  large  quantities  of  vegetables  in  limited  cargo  space.  Clarence 
V.  Ekroth  has  presented  an  excellent  account  of  the  methods  and  Uterature  of  dry- 
ing and  dehydrating  foods  in  Allan  Rogers'  "  Manual  of  Industrial  Chemistry," 
Van  Nostrand,  1918.  The  products  dried  by  Ekroth's  "  G.  H."  Evaporator  in 
the  Mrs.  Oliver  Harriman's  Food  Research  Laboratory  are  of  excellent  quaUty; 
are  said  to  retain  the  important  food  accessories.  The  "  G.  H."  Dehydrator  dries 
its  product  in  moist  air  at  a  moderate  temperature.  Humidity  and  temperature 
are  controlled.  A  fan  blower  is  provided  for  recirculation  of  the  air  so  that 
volatile  substances  are  kept  in  contact  with  the  product  and  only  a  small  per- 
centage is  lost.  The  process  is  quite  the  reverse  of  ordinary  cooking  when  an 
effort  is  made  to  retain  moisture  and  volatile  substances  by  rapidly  seahng  the 
surface  of  the  food  by  quick  heat  coagulation.     Tr.] 


170 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


box.  In  the  first  instance  it  absorbs  approximately  10  per  cent  of  its 
weight  in  the  first  25  hours,  then  shrinking  occurs.  The  ice  box 
heart,  on  the  contrary,  immediately  begins  to  lose  water,  and  at  the 


20 


4.0 


50 


60 


30 
Hours 
Swelling  of  Horse  Heoir+  3or4  Hours, Post  Mortem 

Fig.  a. 

end  of  45  hours  has  lost  by  shrinkage  about  55-75  per  cent  of  its 
water. 

Typical   are  curves,   Figs.   C,  D,  E,   which  show  comparatively 


zo 

tlO 


20 


«51 

c    30 


U    40 


50 


60 


~ 

■" 

~" 

— 

— 

— 

r] 

~ 

— 

~ 

— 

— 

~ 

~ 

^a 

L^ 

\ 

V 

\ 

V 

s 

N 

\ 

\ 

\ 

> 

N 

\ 

*v 

^ 

V 

■»^ 

\ 

^ 

V 

^ 

~> 

> 

y 

\ 

- 

"" 

— 

— 

,.^ 

■- 

— 

._ 

0 

1 

0 

£ 

0 

3 

0 

A 

0 

5 

0 

6 

0 

Hours 

Swelling  of  Horse  Heart  after  Cold  Storage  for 
3  or  4  Hours 

Fig.  B. 


butcher's  meat,  cold  storage  meat  and  hare,  which  have  been  kept  a 
year  at  —  10°.  In  actual  practice  the  method  is  to  weigh  morsels  of 
meat  of  as  nearly  the  same  size  as  possible,  and  after  placing  them  in 
salt  solution,  at  hourly  intervals,  to  determine  the  changes  in  weight. 


FOODS  AND  CONDIMENTS 


171 


Percentage  Change  in  Weight 

I  +  _ 

uit^wro      —      o      —      row*-      ui     g«      -^     o      <o      Q 


o 

X 

■-- 

._^ 

*^ 

"--^ 

*!?•' 

TO 

o 

— 

-^ 

-TOl 

o 

f 

O 

ro 

^ 

^^ 

Ü1 

/ 

Butchers  Meat  (Beef) 
Fig.  C. 


Percentage  Change  in  Weight 


o 

J— w 

" 

—- 



\ 

0 

rj 

^ 

0 

ro 

.^^ 

^'• 

v^ 

/' 

Cold  Storage  Meat 
Fig.  D. 


tn      fi.      (ji      ro 


Percentage  Change  in  Weight 

—        O        —        rOoJ*'<JiS>^0><^0 


0 

-x^ 

'~~~ 



h- 

— 

0 

3 

v-^ 

^X" 

\ 

0 

^ 

y 
0 

/ 

Swelling  of  Hare  Flesh  after  Preservation  for  more  than  a 
Year  at  -10°  C 

Fig.  E. 


172  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Naturally  a  salt  solution  of  the  same  concentration  (5-10  per  cent) 
is  always  employed.  In  pure  water,  shrinking  occurs  immediately, 
since  muscle  albumin  coagulates  in  it  spontaneously.  High  concen- 
trations of  salt,  25-30  per  cent,  likewise  depress  the  curve. 

New  problems  arise  in  cooking  meat.  If  meat  is  boiled  in  pure 
water  we  obtain  a  "weak"  broth.  Muscle  albumin  coagulates  in 
water  before  heat  coagulation  occurs  and  this  impedes  the  exit  of  the 
crystalloid.  Salt  is  therefore  added  immediately  if  good  soup  is  to 
be  expected.  With  hoiling,  heat  coagulation  occurs  whereby  the 
meat  loses  from  20  to  30  per  cent  of  its  water.  The  mechanism  of 
this  loss  is  still  unknown.  We  can  understand  why  there  should  be 
a  loss  of  from  20  to  35  per  cent  in  roasting,  and  it  would  be  still 
greater  if  the  surface  were  not  constantly  protected  by  pouring  over 
or  dipping  into  fat  (basting). 

Preserved  meats  are  less  perishable  because  they  contain  less  water 
and  because  the  muscle  albumin  has  been  converted  into  a  character- 
istic gel  condition.  This  end  is  attained  in  various  ways:  In  pick- 
ling, water  is  removed  from  the  meat  by  the  salts  of  the  brine, 
while  at  the  same  time  there  is  an  exchange  of  crystalloids,  whereby 
salts  enter  from  without  which  change  the  albumin  as  regards  its 
coagulability  and  sweUing  capacity;  and  extractives  leave  it  and 
are  removed  with  the  brine.  Of  course  very  important  changes 
occur  during  storage,  so  that  according  to  A.  Gärtner,  with  in- 
creasing age  pickled  meat  becomes  more  difficult  of  digestion  and 
loses  30  per  cent  of  its  nutritive  value.  Smoking  of  meat  is  usually 
preceded  by  a  short  pickling  process.  The  abstraction  of  water  in 
this  case  is  accomplished  by  means  of  a  strong  current  of  air,  and  in 
the  dried  meat  (pemmican)  which  is  much  rehshed  in  some  regions,  in 
the  Arctics  (for  instance) ,  there  is  no  loss  other  than  water. 

Naturally,  the  properties  of  every  gel,  materially,  depend  upon  its 
history.  To  quote  a  single  example :  F.  Stoffel*  (in  the  laboratory 
of  Prof.  H.  Zangger)  found,  that  the  diffusibility  of  one  and  the 
same  substance  through  the  identical  gelatin  differed,  depending 
upon  whether  the  gelatin  was  rapidly  solidified  with  ice  or  slowly 
cooled  at  room  temperature.  Accordingly,  we  may  assume  in  the 
case  of  meat,  that  the  properties  of  the  coagulated  albumin  will  vary 
with  the  conditions  maintained  during  coagulation,  and  that  upon 
these  depends  its  food  value. 

An  essential  question  in  the  investigation  of  sound  meat,  pre- 
served meats  and  food  preparations  must  be  their  available  food 
value,  which  can  be  answered  only  by  complicated  and  expensive 
metabolism  investigations.  From  my  point  of  view  this  ought  to 
be  a  fruitful  field  for  the  colloid  chemist,  who  ought  certainly  to  be 


FOODS   AND  CONDIMENTS  173 

in  a  position  to  replace  with  simpler  methods  some  protracted  metab- 
olism experiments.  I  might  incidentally  mention  the  methods  of 
adsorption  and  staining  which  hitherto  have  not  been  sufficiently 
considered.  In  the  various  food  preparations  (whose  names  need  not 
be  mentioned),  it  is  quite  unessential  whether  they  contain  a  few  per 
cent  more  or  less  of  carbohydrate  or  nitrogen,  a  fact  which  is  always 
especially  emphasized  in  the  advertisements;  whereas  it  is  quite 
important  to  know  their  swelling  capacity,  and  whether  this  permits 
their  complete  and  rapid  utilization  in  the  alimentary  canal. 

Milk  and  Dairy  Products.  Milk,  as  a  physiological  excretion  will 
be  considered  on  pages  345,  et  seq.,  and  there  also  much  is  said  which 
pertains  to  its  properties  as  a  food  material.  Here  we  shall  concern 
ourselves  merely  with  the  examination  of  milk.  The  present-day 
methods  of  milk  examination  are  limited  to  certain  characteristics 
which  are  especially  easy  to  determine  and,  therefore,  are  easily 
simulated  by  adulterators.  Pure  food  officials  lay  most  stress  on 
the  water  and  the  fat  content.  Sometimes,  in  addition,  they  de- 
termine the  protein  percentage,  preservatives,  and  the  possible  ex- 
istence of  disease  organisms.  Inasmuch  as  milk  is  by  far  the  most 
valuable  food-stuff,  it  is  of  the  greatest  importance  not  only  to 
determine  variations  produced  under  the  normal  circumstances  by 
adulteration,  but  also  those  occurring  under  normal  conditions  of 
production,  change  of  fodder  dependent  upon  change  of  season, 
natural  and  artificial  fodder,  boiling,  pasteurization,  etc.  Accord- 
ingly, H.  Zangger*^  and  his  pupils  undertook  to  discover  new 
methods;  in  them  he  regarded  milk  as  a  solution  of  colloids  and 
electrolytes  Of  the  colloid  methods,  Zangger  and  his  pupil, 
KoBLER,  have  chosen  the  determination  of  surface  tension,  which 
proved  to  be  one  of  the  "most  complicated  but  perhaps  the  most 
deUcate  and  flexible  method."  Among  the  various  procedures,  the 
bubble  method,  in  which  bubbles  are  allowed  to  form  in  the  fluid, 
gave  the  most  constant  results. 

Normal  milk  gave  quite  constant  figures.  Inasmuch  as  by  this 
method  only  such  substances  have  an  influence  as  are  forced  to 
the  surface,  these  can  make  themselves  evident  in  the  minutest 
quantities.  Adulteration  with  water  is  not  easily  detected  by  this 
method.  Fermentation,  on  the  contrary,  causes  great  departures 
from  the  normal,  which  are  explained  by  the  development  of  fatty 
acids.     The  addition  of  alkalis  also  changes  the  surface  tension. 

By  the  study  of  the  viscosity,  abnormal  protein  and  fat  content 
could  be  sho-s\Ti  and  likewise  additions  (adulterations)  which  in- 
fluenced the  amount  of  swelling  (especially  alkaline  additions) .  The 
viscosity  is  also  diminished  by  violent  shaking,  though  milk  regains 


174  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

through  quiet  its  original  viscosity  (to  within  1  per  cent)  provided 
it  has  not  been  shaken  long  enough  for  curds  to  form. 

This  observation  was  of  great  practical  importance  because  milk 
suffers  violent  shaking  during  transportation.  Experiments  in  which 
milk  was  carried  by  wagon,  train  and  post  more  than  three  hundred 
(300)  kilometers  showed  that  there  was  no  evident  irreversible  loss 
of  viscosity. 

Dr.  Grosser,  according  to  a  personal  unpublished  communication, 
has  made  a  very  noteworthy  observation  in  the  ultrafiltration  of 
milk}  It  was  shown  that  raw  milk  gave  an  ultrafiltrate  much 
richer  in  lime  than  did  boiled  milk.  In  boiling,  the  calcium  is  bound 
to  the  milk  colloid  and  remains  with  the  latter  on  the  ultra-filter. 
Thus,  a  simple  means  is  furnished  for  distinguishing  raw  from  cooked 
milk.  Definite  differences  exist  between  human  and  cow's  milk 
which  offer  a  new  basis  for  the  difference  these  two  kinds  of  milk 
exhibit  in  respect  to  their  assimilibility  (available  food  value). 

The  classification  of  the  milk  colloids  to  which  J.  Alexander  and 
J.  G.  M.  BuLLOWA  have  drawn  attention  must  be  considered  in 
future  tests  of  milk  (see  p.  349). 

Since  the  water  and  the  crystalloid  content  of  milk  are  almost  con- 
stant, many  adulterants  can  be  detected  by  the  departure  of  the 
water  and  the  milk  content  from  the  normal.  For  this  purpose  it  is 
necessary  to  remove  the  fat  and  colloid  constituents  without  chang- 
ing the  content  in  water  and  salts.  To  determine  the  addition  of 
water,  J,  Mai  and  S.  Rothenfusser*  coagulate  the  milk  colloids  with 
calcium  chlorid  and  then  measure  the  water  content  by  refraction. 
Kurt  Oppenheimer  determines  the  milk  sugar  polarimetrically,  after 
he  has  removed  the  milk  colloids  with  colloidal  ferric  hydroxid. 
According  to  S.  Rothenfusser,  by  treating  milk  with  lead  acetate 
in  strong  ammoniacal  solution  at  85°  C,  the  milk  sugar  is  adsorbed 
when  the  colloids  are  coagulated,  while  saccharose  remains  in  solu- 
tion. According  to  Rothenfusser,*  the  smallest  adulteration  with 
foreign  sugar  (saccharated  lime)  may  thus  be  detected. 

Among  dairy  products,  condensed  milk  must  be  considered  as  of 
great  importance.  This  is  milk  which  is  evaporated  with  the  addition 
of  25  to  50  per  cent  cane  sugar.  All  who  are  forced  to  use  it,  es- 
pecially colonists,  know  how  ill  it  satisfies  the  demand  for  a  milk 
substitute.  One  of  the  essential  properties  of  colloids  is  that  their 
condition  is  not  reversible  to  the  same  extent  as  crystalloids.  This 
may  be,  in  addition  to  the  destruction  of  certain  flavoring  substances, 
an  important  reason  for  the  lessened  value  of  condensed  milk.  The 
various  dried  milk  products  when  stirred  with  cold  or  warm  water 
^  In  a  private  communication,  as  yet  unpublished. 


FOODS  AND  CONDIMENTS  175 

give  an  incomplete  emulsion  and  tiiere  always  «remains  a  sediment. 
The  older  the  preparation  the  more  incomplete  is  the  solution.  We 
here  approach  once  more  a  phenomenon  which  was  touched  upon  under 
"Aging  of  Colloids"  (p.  74).  J.  G.  M.  Bullowa  informs  me  that 
Just  and  Hatma  ktcr  have  invented  a  process  which  avoids  these 
disadvantages  and  which  is  already  in  use  on  a  large  scale.^ 

Cream  is  a  fat  emulsion  which  contains  at  least  10.  per  cent  fat. 
Cream  for  whipping  contains  as  much  as  30  per  cent.  This  emulsion 
has  the  property  of  building  thick  foam  walls  which  possess  con- 
siderable consistency.  In  order  to  simulate  a  high  fat  content, 
potato  flour,  gelatin  or  whipped  white  of  egg  are  added  as  adulter- 
ants to  cream  deficient  in  fat.  Calcium  saccharate  may  also  raise 
the  viscosity.  S.  M.  Babcock  and  H.  L.  Rüssel*  recommended  its 
addition  to  milk  or  cream  wliich  has  become  thin  from  being  heated. 
The  food  industry  has  adopted  this  and,  nowadays,  calcium  sac- 
charate solutions  enber  conmaerce  under  various  names  (grossin,  etc.) 
as  thickeners.  According  to  Fr.  Elsner,  their  effect  is  quite  mar- 
velous. Their  detection  is  easy  by  the  method  of  Rothenfusser, 
described  on  page  174.  An  artificial  cream  may  be  obtained  by 
emulsifying  warm  margarine  with  skim-milk  and  adding  egg  yolk. 

It  is  evident  fiom  what  was  said  on  pages  15  and  34,  why  an 
emulsion  such  as  whipped  cream  or  the  like  is  so  stiff,  because  we 
know  how  great  a  force  is  necessary  to  deform  spheres  of  such  small 
size. 

In  milk  and  cream,  the  aqueous  colloid  solution  is  the  dispersing 
medium  and  the  fat  is  the  dispersed  phase;  in  the  case  of  butter  this 
relation  is  reversed.^  According  to  law,  butter  may  not  contain 
more  than  16  per  cent  water,  though  it  is  possible  to  impregnate 
it  with  water  to  more  than  30  per  cent."  According  to  Posnjak, 
the  addition  of  alkalis  and  glucose  increases,  whereas  increase  of 
acidity  diminishes  the  capacity  of  butter  to  absorb  water.  (W. 
Meijeringh.*)  The  kneading  in  of  water  is  always  reckoned  to 
be  an  adulteration,  because  water  is  cheaper  than  butter.  From 
the  standpoint  of  the  colloid  chemist,  it  has  always  been  a  question 
whether  the  amount  of  water  in  butter  or  rather  the  content  of  skim- 
milk  does  not  increase  its  digestibility  and  whether  it  is  not  the 
dispersion  by  means  of  the  albumin  or  rather  casern-containing 
aqueous  solution  which  makes  butter  so  much  superior  in  digesti- 
bility to  other  fats  of  high  melting  point;  and  whether,  if  the  above 
assumption  should  be  proved  correct,  it  would  not  be  possible  to 
find  a  legal  way  to  permit  butter  to  have  a  greater  water  (i.e.,  skim- 
milk)    content.     In   the   manufacture   of   margarine,    skim-milk   is 

1  [Merrall  and  Soule  of  Rochester,  N.  Y.,  spray  milk  into  heated  air  to 
dry  it.    Tr.] 

2  [See  work  of  Martin  H.  Fischer  and  G.  F.  L.  Clowes.     Tr.] 


176  COLLOIDS  IN  BIOLOGY  AND   MEDICINE 

added  to  the  fat  in  order,  we  are  accustomed  to  assume,  to  give  it 
the  taste  of  butter.  It  is  yet  to  be  determined  whether  it  is  not  just 
this  addition,  which  gives  the  fat  the  dispersion  characteristic  of 
butter,  and  thereby  its  greater  digestibility.  The  darkening  of 
margarine  in  heating  is  undoubtedly  to  be  attributed  to  the  added 
skim-milk. 

By  changing  the  colloidally  dissolved  albuminous  substances  of 
milk  into  the  gel  form,  we  obtain  cheese.  Coagulation  can  be  brought 
about  by  means  of  rennet  (sweet  milk  cheese)  or  through  acidification 
(sour  milk  cheese).  In  cheese  we  have  an  emulsion  of  fat  in  a  protein 
gel;  whereas  in  skim  or  sour  milk  cheese  (kümmel,  Harz  and  hand- 
käse), the  amount  of  fat  is  only  the  small  amount  afforded  by  skim- 
milk;  it  is  quite  high  in  the  fatty  cheeses  (cream,  Swiss,  Camembert 
and  Roquefort). 

A  process  of  great  interest,  as  yet  uninvestigated  from  the  colloid- 
chemical  standpoint  is  the  ripening  of  cheese.  Through  the  action 
of  bacteria  there  occur  changes  in  the  structure  of  the  cheese  which 
are  specific  for  every  variety  and  which  cause  the  spotted  appear- 
ance found  in  the  various  kinds  of  cheese. 

For  cheese,  chemical  tests  are  limited  to  the  determination  of  the 
water,  fat,  albumin  and  salt  content  and  the  possible  adulterants. 
The  most  important,  namely,  the  swelling  capacity  in  the  presence 
of  the  digestive  ferments,  is  nowadays  entirely  ignored,  although 
this  would  furnish  the  simplest  method  of  deciding  the  important 
question  of  the  digestibility  of  cheese. 

Honey  should  in  the  main  be  composed  of  sugar;  it  is  neverthe- 
less frequently  adulterated  with  glucose  and  dextrin.  My  opinion 
is  that  tests  of  the  surface  tension  of  dilute  solutions  would  lead  to 
the  detection  of  such  colloidal  adulterants. 

Flour,  Dough  and  Baking  Products. 

The  examination  of  flour,  in  addition  to  the  microscopic  histologic 
study,  extends  to  its  doughing  and  baking  properties.  These  two 
questions  belong  entirely  to  the  province  of  colloid  chemistry. 

Art  is  in  advance  of  science  in  this  matter.  There  exist  the  most 
diverse  methods  for  discovering  the  presence  of  foreign  substances 
in  flour  and  for  distinguishing  its  various  varieties  by  determining 
the  temperature  at  which  it  becomes  pasty.  The  baking  capacity 
in  particular,  which  is  intimately  bound  up  with  the  swelling  capacity 
of  the  gluten,  is  applied  colloid  chemistry.  The  more  glutenous  the 
flour,  the  more  water  it  binds  (38  to  60  per  cent)  and  the  greater  is 
its  capacity  to  be  kneaded. 


FOODS  AND  CONDIMENTS  177 

The  apparatus  for  this  determination,  especially  that  of  Leo 
Liebermann,  measures  the  expansion  of  the  "doughed  up"  flour 
under  the  influence  of  heat. 

I  must  not  omit  to  state  here,  that  flour  whose  baking  capacity- 
has  suffered  (for  instance  by  over-heathig  the  gluten  in  grinding) 
may  be  restored  by  the  addition  of  common  salt,  plaster  of  paris, 
water  and  alum. 

What  has  been  said  of  flour  applies  also  to  prepared  flours  and  in- 
fant foods.  In  the  latter,  in  addition  to  the  proper  composition,  ease 
of  digestion  and  the  property  of  preventing  curdy  coagulation  of 
milk  in  the  stomach  must  be  considered.  It  should  be  determined 
whether  a  part  of  the  difficult  and  complicated  metabolism  ex- 
periments could  not  be  substituted  by  simple  testing  according  to 
suitable  colloid-chemical  methods  (swelling,  etc.). 

In  the  investigation  of  dough,  egg-dough  and  prepared  products 
(bread,  noodles  and  macaroni),  there  occurs  a  phenomenon  which  is 
very  suggestive  of  a  similar  occurrence  in  the  case  of  milk  (see  p. 
345),  namely,  that  one  cannot  recover  the  quantity  of  fat  present  in 
the  original  flour  by  means  of  ether  extraction,  and  indeed,  it  would 
be  interesting  to  determine  how  the  adsorption  of  the  fat  occurs  if 
there  is  any  adsorption.  In  this  connection  we  may  consider  that  in 
the  case  of  products  made  of  egg-dough,  we  distinguish  between  free 
lecithin  (extractible  with  ether)  and  hound  lecithin  (extractible  with 
alcohol).     Perhaps  here,  too,  it  is  a  question  of  adsorption. 

Next  to  milk,  bread  is  our  most  important  foodstuff.  Bread  making 
may  be  briefly  mentioned  here.  Bread  is  prepared  from  flour,  which, 
if  it  were  consumed  directly  or  made  into  a  paste,  would  be  badly 
digested  because  the  flour  grains  possess  only  a  small  swelling  capacity 
and  the  surface  development  of  the  entire  mass  is  very  small.  The 
making  of  bread  renders  the  individual  parts  easily  accessible  to  the 
digestive  juices.  For  this  purpose,  the  dough  (flour  mixed  with  water) 
is  caused  to  ferment  by  the  addition  of  yeast  or  sour  dough.'  As  a 
result,  the  starch  grains  swell,  burst  and  take  up  water,  a  portion  is 
converted  into  dextrin,  part  of  which  is  still  further  broken  down 
into  sugar,  alcohol  and  carbonic  acid  gas.  By  foam  formation, 
the  carbonic  acid  gas  causes  an  enormous  increase  in  the  surface  of 
the  mass.  Incidental  fermentative  processes  give  the  gluten,  the 
plant  albumin,  the  power  to  swell  up.  This  condition  is  completed 
and  to  a  certain  extent  fixed  by  baking.  The  dextrinization  of  the 
starch  is  thus  completed,  the  development  of  surface  is  increased  b}^ 
the  conversion  of  the  water  into  steam  and  the  expansion  of  the 
carbonic  acid  gas,  the  gluten  is  coagulated  and  further  changes  are 
stopped  by  the  killing  of  the  fermenting  agents. 


178  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Ultimately  we  obtain  a  framework  of  coagulated  gluten  whose 
pores  are  filled  with  shattered  starch  grains. 

The  crust,  which  does  not  swell  much,  acts  as  a  protection 
both  against  the  absorption  of  water  and  its  loss  from  the  interior. 
A  good  bread  should  contain  from  35  to  45  per  cent  of  water. 
Upon  keeping,  it  loses  about  1  per  cent  daily  until  the  loss  reaches 
15  per  cent.  After  that,  the  water  content  is  dependent  on  the 
humidity  of  the  atmosphere;  this  corresponds  to  the  behavior  of  an 
elastic  gel.  It  is  interesting  to  note  that  the  salt  content  plays  a 
role  in  the  condition  of  swelling,  because  unsalted  bread  dries  much 
more  readily  than  salted  bread. 

There  is  a  widespread  error  that  stale  bread  has  lost  water  and  is 
dessicated.  This  is  not  true;  the  crumbling  consistence  of  stale 
bread  is  due  to  a  shifting  of  the  water  within  the  loaf;  the  starch 
grains  transfer  water  to  the  albuminous  framework.  J.  R.  Katz 
studied  this  problem  and  found  that  bread  kept  fresh  longer  at  50°  to 
100°  C.  as  well  as  below  — 10°  (best  in  a  current  of  air),  in  other  words, 
there  is  a  balance  of  swelling  in  starch  and  gluten  which  corresponds 
to  that  of  fresh  bread.  At  from  0°-25°  C.  stale  bread  is  the  stable 
form.  Staling  is  a  particularly  reversible  process;  dry  rolls  are  made 
fresh  by  heating  them.  This  is  an  old  expedient  frequently  employed. 
The  results  of  Katz'  research  on  keeping  bread  fresh  at  low  tem- 
peratures deserves  the  attention  of  the  trade. 

The  digestibility  and  available  food  value  of  bread  depend  par- 
ticularly upon  its  "  dispersibility  "  and  swelling  capacity. 

A  perfect  wheat  bread  may  be  utilized  to  the  extent  of  94  per  cent 
—  a  rye  bread  to  90  per  cent.  For  this  purpose,  the  flour  used  must 
be  as  fine  as  possible,  otherwise  the  utilization  is  imperfect.  It  must 
also  be  properly  swollen  up;  fresh  bread  which  is  too  wet  is  digestible 
with  difficulty.  It  packs  together  and  the  changes  induced  by  the 
incorporation  with  the  saliva  and  other  digestive  juices  are  differ- 
ent from  those  with  old  or  dry  bread.  Most  difficult  to  absorb  are 
the  proteins  (55  to  85  per  cent).  The  great  heat  (in  the  inner  parts 
amounting  to  110°),  acting  in  the  presence  of  a  small  quantity  of 
water,  produces  a  coagulation  which  greatly  reduces  their  swelling 
capacity. 

The  grain  shortage  during  the  war  caused  the  attempt  to  make 
bread  from  potatoes.  Potato  flour  yields  a  heavy,  indigestible  mass 
when  heated  in  the  way  usual  for  ordinary  flour.  Different  attempts 
were  made  to  overcome  this.  According  to  A.  Fornet,  the  Ex- 
perimental Station  for  the  Utilization  of  Grain  mixes  in  an  unknown 
gluten  substitute.  Wilhelm  Ostwald  recommended  blood  or 
casein  dissolved  in  ammonium  carbonate  as  a  substitute  for  gluten. 


FOODS  AND  CONDIMENTS  179 

Walter  Ostwald  and  A.  Riedel  made  a  porous  starch  bread  by 
adding  a  starch  paste  to  the  starch  dough  before  baking.  A  ''  pseudo- 
coagulation"  occurred  during  the  baking,  the  unburst  starch  grains 
abstracting  water  from  the  burst  ones.  The  internal  friction  of  the 
paste  becomes  so  great  that  the  air  bubbles  cannot  escape  during 
the  baking  process,  the  dough  does  not  fall  but  is  fixed  as  a  foam. 

[In  discussing  the  physical  chemistry  of  bread  making,  E.  J.  Cohn  and  L.  J. 
Henderson  (Science,  Nov.  22,  1918,  p.  501,  et  seq.)  conckide  that  "the  acidity  of 
the  dough,  at  the  time  of  baking,  seems  to  be  the  most  important  variable  factor 
in  bread  making."  Soluble  serum  protein  is  an  acceptable  physical  substitute 
for  gluten.     Tr.] 

Beer. 

The  fermentation  industry  so  highly  developed  scientifically  and 
technically  has  already  paid  attention  to  colloid  chemistry  and  pro- 
duced a  not  inconsiderable  literature  (see  F.  Emslander  *^)  which 
in  part,  however,  is  not  altogether  free  from  dilettantism. 

It  would  take  us  too  far  afield,  were  we  to  consider  the  whole 
process  of  beer  brewing  ^  from  the  colloid-chemical  viewpoint;  we 
must  restrict  ourselves  to  the  finished  product. 

Beer  is  a  fermented  beverage  with  an  alcohol  content  of  from  2  to 
5  per  cent,  some  acetic  acid  and  from  4  to  8  per  cent  extractives. 

The  extractives  consist  in  greatest  part  of  carbohydrates  (maltose, 
dextrin  and  gums),  to  a  lesser  extent  of  proteins  (about  0.6  per  cent), 
and  in  addition,  salts,  hop  bitters,  hop  resin  and  several  alkaloidal 
substances,  besides  small  quantities  of  fermentation  products  such 
as  glycerin,  lactic  acid  and  succinic  acid. 

The  persistent  fine  foam  which  a  fresh  beer  should  show  is  brought 
about  by  its  colloidal  content.  It  is  a  sign  that  the  colloids  have  not 
j^et  been  broken  down  too  far,  and  has  at  the  same  time  the  more 
important  purpose  of  retaining  the  carbonic  acid  gas.  In  a  solution 
supersaturated  with  gases,  the  formation  of  bubbles  is  either  in- 
creased or  diminished  by  the  colloids  present  at  the  moment.  We 
know  further,  from  page  34,  that  a  certain  pressure  is  necessary  to 
overcome  the  surface  tension  and  burst  the  bubble,  e.g.,  a  soap  bubble, 
so  that  the  carbonic  acid  gas  of  beer  is  under  a  certain  pressure 
beneath  the  foam. 

The  condition  rather  than  the  amount  of  the  foam-forming  albu- 
mins is  more  important  for  the  foam-keeping  quality  of  beer.  There 
are  beers  rich  in  albumin  which  remain  foamless  and  beers  poor  in 
albumin  which  foam  well.  According  to  F.  Emslander*  it  is  mainly 
the  soft  hop  resin,  in  addition  to  the  acidity  of  the  beer,  which  makes 
the  albumins  foam. 

1  Rich.  Emslander  calls  attention  to  an  interesting  relation  between  beer 
brewing  and  inactivation  of  ferments  by  shaking,  see  p.  189.  Brewers  have  long 
known  that  shaking  by  trains,  machines,  etc.,  interferes  with  the  fermentation 
and  lagering  of  beer. 


180  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

A  perfect  beer  should  be  absolutely  clear;  turbid  beers  are  unre- 
liable, but  no  objection  can  be  raised  to  a  dusty  or  net-like  appearance. 
In  the  latter  case  the  disperse  phase  consists  of  protein  particles,  dex- 
trins  or  precipitates  of  hop  resin.     Some  yeast  may  also  be  suspended. 

Occasionally  in  very  cold  beers  cloudiness  develops  which  may  be 
ascribed  to  precipitated  albumins,  which  disappear  when  the  beer  is 
warmed.  In  the  United  States  where  iced  drinks  are  in  great  de- 
mand, especial  pains  have  been  taken  to  master  the  difficulty. 

The  alkalinity  of  the  wash  water,  the  carbonic  acid,  and  the 
atmospheric  oxygen  during  the  brew,  play  an  important  role  in  the 
resistance  of  beer  to  cold.  According  to  R.  Emslander*  the  surest 
means  is  the  addition  of  some  pepsin.  [Wallerstein  has  patented 
the  addition  of  a  proteolytic  enzyme  to  beers,  to  prevent  cold- 
cloudiness.     Tr.] 

What  is  designated  as  "vollmundigkeit"  or  body  in  beer  is  caused 
by  the  colloid  content.  This  property  is  almost  identical  with  the 
viscosity,  and  is  determined  by  the  viscosimeter.  If,  for  instance,  the 
time  required  for  water  to  run  from  a  50-c.c  pipette  =  1,  and  that  of 
an  equal  quantity  of  beer  =  1.43,  we  say  that  the  "viscosity"  is  1.43. 
It  follows  from  what  has  been  said  on  pages  152  and  153,  that  we 
may  assume  d  priori,  that  the  "vollmundigkeit"  is  largely  dependent 
on  the  electrolytes,  that  is,  on  the  content  of  acids  and  the  kinds  of  salts. 
Even  though  the  larger  part  of  the  salts  is  derived  from  barley,  yet 
some  are  derived  from  the  brewing  water,  and  the  hitherto  partly 
unrecognized  influence  of  the  water  may  be  attributed  to  this  fact. 

E.  MouFANG  determined  empirically  the  relation  between  optimum 
keeping  quality,  "full"  and  "palatable"  taste,  sediment  and  acidity. 
I  refer  to  F.  Emslander  *^  for  the  colloid-chemical  effect  of  the 
brewing  water  on  lagered  beer. 

Among  the  proteins,  in  addition  to  the  gluten  which  flocculates  out 
on  boiling  with  acetic  acid,  peptone  may  be  mentioned.  H.  Bech- 
HOLD  *  ^  was  able  to  demonstrate  only  albumoses,  upon  examining  a 
beer  by  ultrafiltration.  Before  generalizing,  a  large  number  of  beers 
would  have  to  be  investigated  in  this  way.  It  seems  that  E.  Fouard 
has  carried  on  such  ultrafiltration  experiments  with  starch  solutions, 
worts  and  beer  (cited  by  Emslander).  W.  H.  van  Laer  has  also 
made  noteworthy  experiments  on  the  relationship  between  the  ultra- 
filtrates  of  beer  and  musts,  and  their  transparency.  F.  Emslander 
and  H.  Freundlich  *  have  performed  cataphoretic  experiments  and 
found  that  the  colloids  migrate  to  the  cathode.  In  consideration  of 
the  acid  content  of  beer,  this  finding  is  theoretically  correct. 

R.  Marc  *  has  worked  out  a  simple  method  for  quantitatively 
determining  beer  colloids  by  means  of  the  fluid  interferometer. 


FOODS  AND  CONDIMENTS  181 

A  study  should  be  made  of  the  usefulness  of  several  other  colloid- 
chemical  methods  for  the  testing  of  beer,  especially  the  determina- 
tion of  surface  tension,  which  might  serve  to  distinguish  the  amount 
of  various  colloids  contained,  coagulation  methods,  etc.  The  mere 
suggestion  should  suffice. 

It  must  be  mentioned  in  addition,  that  F.  Emslander*^  has  at- 
tributed to  the  "protective  colloids"  of  beer  a  significance  for  the 
more  easy  adsorption  of  milk  and  other  foodstuffs.  Even  earfier 
experiments,  especially  those  of  Ross  van  Lennep  indicate  that  the 
presence  of  colloids  and  suspensions  have  an  influence  on  the  growth 
of  microorganisms.  S'hngen  thoroughly  investigated  this  matter 
in  connection  with  alcoholic  fermentation  and  obtained  interesting 
results.  He  found  that  colloidal  iron,  albumin,  silicic  oxid  and 
humic  acid  had  no  influence  on  alcoholic  fermentation,  but  that,  on 
the  contrary,  it  was  greatly  hastened  by  turf,  filter  paper,  blood 
charcoal  and  garden  earth.  He  succeeded  in  proving  the  cause  of 
this;  the  carbonic  acid  which  is  developed  during  alcoholic  fermen- 
tation impedes  fermentation  and  all  substances  which  favor  the  dis- 
appearance of  the  carbonic  acid  favor  fermentation.  The  action  of 
the  colloids  mentioned  is  purely  mechanical,  somewhat  like  that  of 
powdered  glass,  threads,  wood  chips  or  platinum  shavings  which 
hinder  boiling.  In  the  fermentation  industry  it  is  generally  known 
that  brewers  grains  and  spun  glass  increase  alcohoHc  fermentation, 
and  these  phenomena  have  now  been  explained  by  Söhngen's 
investigation. 


S.  Rothenfusser  *  has  employed  his  colloid-adsorption  method 
for  detecting  saccharose  in  the  most  diverse  kinds  of  foods  and  condi- 
ments (wine,  weissbier,  cafö  parfait,  kilned  malt,  pastry,  etc.). 

In  practice  naturally  many  other  questions  will  appeal  to  food 
chemists.  It  might  be  determined  whether  the  availability  of  vege- 
table protein,  which  on  digestion  is  only  from  60  to  70  per  cent, 
could  not  be  increased  by  a  suitable  method  of  preparation.  Colloid- 
chemical  methods  must  unquestionably  be  utilized  in  the  investiga- 
tion of  fruit  juices,  jellies  and  marmalades.  We  must  remember  that 
these  are  frequently  mixed  with  glucose,  which,  when  undeclared, 
should  be  regarded  as  an  adulteration.  Glucose  contains  in  addition 
to  dextrose,  dextrin  and  unfermentable  substances  which  may  be 
determined  by  colloid-chemical  analysis.  Marmalades  are  adulter- 
ated with  gelatin,  agar-agar  and  isinglass. 

We  trust  that  the  mere  mention  of  these  facts  may  cause  food 
chemists  to  give  greater  attention  to  colloid-chemical  methods. 


CHAPTER  XII. 
ENZYMES. 

For  more  detailed  study  we  recommend  the  following  books  of  reference: 
"The  Nature  of  Enzyme  Action,"  by  W.  M.  Bayliss;  "Allgem.  Chemie  der 
Enzyme,"  by  H.  Euler  (J.  F.  Bergmann,  Wiesbaden,  1910);  "Die  Fermente 
und  ihre  Wirkungen,"  by  C.  Oppenheimer  (F.  C.  W.  Vogel,  Leipzig,  1913); 
["  Biochemical  Catalysis  in  Life  and  Industry,"  by  Jean  Effront.  Translated 
by  Samuel  Prescott.    John  Wiley  &  Sons,  Inc.,  1917.    Tr.] 

List  of  the  best  known  enzymes. 

Amylase  hydrolyzes  starches  and  glycogen  into  dextrin  and  maltose. 

Catalase  decomposes  peroxid  of  hydrogen. 

Chymosin  is  rennin. 

Diastase  fluidifies  starches  and  hydrolyzes  them  to  maltose. 

Emulsin  hydrolyzes  glucosides. 

Erepsin  hydrolyzes  albumoses  and  peptones  to  amino-acids. 

Fibrin-ferment  an  hypothetical  ferment  which  coagulates  fibrin. 

Invertase  hydrolyzes  cane  sugar. 

Lipase  hydrolyzes  fats  into  fatty  acids  and  glycerin. 

Maltase  cleaves  glucosides. 

Oxidase  an  oxygen  carrier. 

Pancreatin  from  the  pancreatic  juice  is  a  mixture  of  several  enzymes. 

Papain  hydrolyzes  albumin. 

Pepsin  hydrolyzes  albumin  in  acid  solution. 

Ptyalin  is  the  amylase  of  the  saUva. 

Rennin  coagulates  milk. 

Steapsin  is  hpase. 

Trypsin  hydrolyzes  albumin  in  alkaHne  solution. 

Tyrosinase  oxidizes  tyrosin  and  some  of  its  derivatives. 

Zymase  splits  sugar  into  alcohol  and  CO2. 

To  split  complex  molecules,  chemists  have  to  employ  powerful  re- 
agents, such  as  acids,  alkahs,  etc.  They  smash,  as  it  were,  the 
clockwork  with  a  hammer  and  then  pick  out  the  undamaged  particles. 
Just  as  a  watchmaker  employs  for  each  screw  a  suitable  tool  or  a 
specially  made  pliers,  so  nature  has  constructed  delicate  instruments 
for  this  purpose.  Enzymes  are  such  tools  for  the  chemical  break- 
ing down  or  building  up  of  molecules.  Albumin,  carbohydrates  and 
fats  may  all  be  split  up  by  acids.  For  each  purpose  nature  has  a 
special  enzyme,  or  even  several;  for  the  cleavage  of  albumin,  pepsin 
and  trypsin;  for  starches,  diastase;  for  fats,  lipase. 

We  shall  see  that  some  enzymes  are  fashioned  exactly  for  their 
use,  so  that  the  simile  of  Emil  Fischer,  which  compares  the  enzyme 

182 


ENZYMES  183 

to  a  key  and  the  substance  split  up  to  a  lock,  is  a  very  happy  one. 
The  simile  can  be  extended  still  further,  since  the  key  may  unlock 
thousands  of  similar  locks  and  fails  only  when  the  key  is  worn  out. 
Moreover,  only  very  small  quantities  of  enzymes  are  needed  which 
are  utihzed  over  and  over  again.  This  conception  of  enzymes  corre- 
sponds with  our  present-day  chemical  conception  of  catalyzers.  These 
latter  are  substances  which  either  bring  about  or  accelerate  chemical 
reactions,  without  themselves  figuring  in  the  end  products.  For  in- 
stance, platinum  hastens  the  combination  of  O  and  SO2  into  SO3,  or 
the  union  of  H2  and  0  into  H2O. 

It  is  the  nature  of  catalyzers  that  they  split  up  compound  sub- 
stances and  build  up  the  same  substances  from  the  cleavage  products 
until  a  definite  equilibrium  is  obtained.  Thus,  ricin,  the  enzyme  of 
the  castor  bean,  not  only  splits  fat  into  glycerin  and  fatty  acid  but 
also  unites  glycerin  and  fatty  acids  into  fats.  The  equihbrium  is 
frequently  one  in  which  the  synthetic  action  seems  very  subordinate. 
Thus,  amylase  under  certain  circumstances  splits  up  99  per  cent 
of  starch,  yet  it  forms  possibly  but  1  per  cent  of  starch  from  maltose. 
This,  being  a  colloid,  precipitates  from  the  solution,  and  thus  permits 
the  formation  of  another  1  per  cent  of  starch  which  gradually  ap- 
pears as  starch  grains  or  glycogen  and  thus  permits  the  further  for- 
mation of  starch. 

Hitherto,  it  has  been  impossible  to  obtain  an  enzyme  in  pure  form 
and  only  by  its  stronger  or  weaker  activity  was  it  known  whether  a 
dilute  or  a  concentrated  preparation  of  enzyme  existed.  W.  M. 
Bayliss  rightly  calls  attention  to  the  fact  that,  on  account  of  their 
colloidal  nature,  enzymes  always  carry  down  by  adsorption  portions 
of  the  solutions  from  which  they  are  obtained.  It  is,  therefore,  not 
surprising  that  albumin  reactions  are  obtained  from  pepsin  and  tryp- 
sin and  a  carbohydrate  reaction  from  amylase  and  invertase.  In 
many  cases  it  is  possible  to  decide  by  means  of  diffusion  whether 
mixtures  are  present  (see  R.  0.  Heezog  and  Kasarnowski*).  Ac- 
cording to  L.  RosENTHALER,*  the  presence  of  albuminous  substances 
is  of  biological  importance,  protecting  the  enzyme  from  many  de- 
structive influences,  especially  from  H  and  OH  ions.  As  the  result 
of  the  constant  presence  of  adsorbed  impurities,  we  know  almost 
nothing  concerning  the  chemical  nature  of  enzymes.  However,  we 
do  know  that  all  enzymes  are  colloids. 

S.  Fränkel  assumes  that  pure  diastase  is  very  greatly  dispersed 
but  as  yet  no  sufficient  evidence  has  been  adduced;  neither  observa- 
tion in  the  ultramicroscope  nor  filtration  through  a  4  per  cent  ultra- 
filter  is  conclusive.  To  us  it  merely  seems  probable  that  such 
diastase  is  more  highly  dispersed  than  hemoglobin. 


184  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Even  the  colloidal  state  itself,  i.e.,  great  surface  development,  under 
certain  circumstances,  may  be  responsible  for  work  similar  to  that  per- 
formed by  certain  of  the  enzymes.  For  instance,  G.  Bredig  catalyzed 
hydrogen  peroxid  by  means  of  metal  sols,  particularly  platinum  sol, 
which  he  prepared  by  electric  pulverization;  that  is,  he  obtained  a 
result,  the  splitting  off  of  oxygen,  which  in  all  appearances  resembles 
that  brought  about  by  catalase,  a  ferment  which  occurs  in  blood,  in 
milk,  and  in  many  plant  and  animal  tissues.  On  this  account  G. 
Bredig  called  his  metal  sols  "inorganic  ferments,"  although  with 
enzymes  (or  ferments),  they  share  other  properties  to  which  we  shall 
return.  The  action  of  enzymes  is  explained  in  part  by  their  colloidal 
nature.  In  the  organism  they  act  chiefly  on  colloid  substances 
(e.g.,  foods)  with  very  extensively  developed  surfaces,  so  that  under 
certain  circumstances  enzymes  may  be  merely  mechanically  adsorbed. 
They  consequently  act  upon  the  substrate  in  the  greatest  concentra- 
tion. 

It  was  shown  by  numerous  adsorption  experiments  with  indiffer- 
ent suspensions  (charcoal,  kaolin,  cellulose)  that  enzymes  have  a 
strong  tendency  to  concentrate  on  surfaces.  It  is  possible  to  remove 
the  rennin  or  pepsin  (M.  Jacoby*),  and  trypsin  (G.  Büchner  and 
Klatte*)  from  solution  by  means  of  fibrin  flakes  or  other  coagulated 
albumin,  or  diastase  by  means  of  starch  (H.  van  Laer).  The 
reagents  and  sometimes  also  the  products  of  the  reaction  are  adsorbed 
by  the  colloidal  enzymes.  If  the  former  accumulate,  in  accordance 
with  recognized  laws,  the  progress  of  the  reaction  is  slowed.  An 
example  is  the  breaking  down  of  hydrogen  peroxid  by  catalase.  The 
oxygen  formed  by  the  breaking  down  of  H2O2  into  H2O  and  O  is  ad- 
sorbed by  catalase  and  the  reaction  is  slowed  (Waentig  and  Steche). 
Some  enzymes,  especially  pepsin  and  papagotin,  according  to  Rohongi, 
give  reversible  precipitates  in  salt-free,  neutral  solutions  of  different 
albumins  on  which  they  act.  The  inhibition  of  the  action  of  an 
enzyme  by  a  suspension  or  a  colloid  may  be  removed  again  under 
certain  conditions  by  another  indifferent  colloid.  If  the  activity  of 
rennet  has  been  destroyed  by  charcoal  or  normal  serum  so  that  the 
mixture  no  longer  produces  curdling  of  milk,  the  activity  of  the 
rennet  may  be  restored  by  the  addition  of  saponin.  Somewhat 
different  modifications  are  obtained  by  the  addition  of  Cholesterin 
or  by  combinations  of  trypsin  with  charcoal,  saponin  or  Cholesterin 
(Johnson,  Blüm*).  In  this  way,  the  numerous  possibilities  which 
result  from  the  interaction  of  enzyme  and  antienzyme  (q.v.)  rest  on 
a  physical  basis. 

The  essential  difference  between  an  indigestible  adsorbent  and  one 
which  is  dissolved  by  the  enzyme  is  that  the  combination,  e.g.,  be- 


ENZYMES  185 

tween  animal  charcoal  and  trypsin,  is  primarily  irreversible.  The 
trypsin  is  fixed,  water  is  unable  to  tear  the  trypsin  from  the  char- 
coal though  casein  is  able  to  do  so  (S.  G.  Hedin).  It  is  seen,  accord- 
ingly, that  trypsin  undergoes  a  change  on  the  surface  of  charcoal 
similar  to  that  undergone  by  dyes  on  fibers  and  if  the  process  were 
to  occur  in  the  organism  as  it  does  on  charcoal,  the  trypsin  would 
be  permanently  withdrawn  from  the  mixture.  If,  however,  the  sub- 
strate is  digested  and  crystalloid  cleavage  products  result,  e.g.,  in  the 
cleavage  of  fibrin  by  trypsin,  the  adsorption  ceases  of  its  own  accord 
and  the  enzyme  becomes  free  for  further  use,  acting  like  a  true 
catalyzer. 

This  serves  to  explain  the  significance  of  the  surfaces  of  the  sub- 
strate on  enzyme  action.  The  action  increases  in  speed  in  accord- 
ance with  extent  of  surface  per  unit  of  weight.  E.  Abderhalden 
and  Pettibone*  demonstrated  this  in  the  digestion  with  pancreatic 
juice  of  albumen  coagulated  in  different  ways. 

In  the  case  of  enzymes  their  electrochemical  nature  is  more  im- 
portant than  in  the  case  of  other  colloids,  and  influences  their  ad- 
sorption. 

We  frequently  observe  that  if  the  proper  H  or  OH  ion  concentra- 
tion is  absent,  an  enzyme  acts  feebly  on  its  substrate.  Many  neutral 
salts  favor  enzyme  action;  others  inhibit  it.  For  instance,  pepsin 
acts  strongly  in  acid  solution  only,  trypsin  in  alkaline  solution  only. 

The  investigations  of  L.  Michaelis*^  and  his  co-workers  show 
that  the  electric  charge  of  different  enzymes  varies  (see  H.  Iscov- 
Esco*^)  and  that,  proportionately  to  the  charge,  they  are  unequally 
adsorbed  by  various  substrates. 

We  have  previously  mentioned  that  electropositive  gels  or  suspen- 
sions, e.g.,  ferric  oxid,  completely  adsorb  electronegative  solutions, 
e.g.,  serum  albumin.  An  electronegatively-charged  mastic  or  kaolin 
suspension  completely  attracts  to  itself  serum  albumin,  only  when  it 
has  become  electropositive  by  acidification. 

These  investigations  gave  the  following  results  for  a  group  of 
enzymes.  In  the  table  (see  p.  186)  X  signifies  a  pronounced  electric 
migration  (to  cathode  or  anode),  i.e.,  complete  adsorption;  0  signi- 
fies no  migration  or  no  adsorption;  X  —  0,  0  —  X,  respectively, 
more  or  less  migration  or  adsorption. 


186 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Migration  towards 

Adsorbents 

Ferment. 

Anode 

+ 

Cathode 

+ 

Iron  oxid, 
clay,  etc. 

Kaolin, 

mastic, 

arsenious 

sulphid, 

etc. 

neutral 
charcoal. 

Invertase: 

in  neutral  solution 

in  acid,  solution.  .  . 

X 
X 

0-  X 
0 
X 

X 
0 

X 

X 

X 

destroyed 

0 
0 

X 

X  -0 
0 

0 
X 
0 

0 

0 

destroyed 

X 
X 
X 

X 

0-  X 
X 

X 

X 
X 

X 

X 

X  -0 

X 

X 

destroyed 

0 
0 
0 

0 
X 
0 

X 
X 
X 

X 

X 

0-  X 

0 
X 

destroyed 

0 
0 

destroyed 

X 
destroyed 

X 

X 

in  alkaline  solution 

Plant  diastase: 

in  neutral  solution 

in  acid  solution 

X 

X 
X 

in  alkaline  solution 

Salivary  diastase: 

in  neutral  solution 

in  acid  solution 

0 

X 
X 

in  alkaline  solution 

Trypsin: 

in  neutral  solution 

in  acid  solution 

X 

X  -0 
X 

in  alkaline  solution 

Pepsin: 

in  neutral  solution. 

in  acid  solution 

0-  X 

X 
X 

in  alkaline  solution 

Rennin  (from  pepsin) : 

in  neutral  solution 

in  acid  solution 

destroyed 

in  alkaline  solution 

Rennin  (Grübler): 

in  neutral  solution 

in  acid  solution 

in  alkaline  solution 

We  learn  from  this  table  that  analysis  by  simple  adsorption  may 
replace  the  more  difficult  and  complicated  electric  migration.  It  is 
very  instructive  for  our  knowledge  of  enzymes,  that  their  action  is 
strongest  with  the  reaction  which  expresses  their  own  charges  as  may 
be  seen  from  the  following  table  (from  L.  Michaelis)  : 

Optimum  Activity  Occurs 
with  an  H-ion  Concentration  of 

Water 1 .  10-^ 

Invertase 2 .  10~^ 

Maltase 2.5.10-^ 

Trypsin 2. 10^ 

Erepsin 2 .  10-^ 

Pancreatic  lipase 2 .  10~^ 

Pepsin 2. 10-2 


ENZYMES  187 

Electronegative  (acid) "pepsin  digests  best  in  an  acid;  and  ampho- 
teric (almost  neutral)  trypsin  in  an  alkaline  reaction.  We  may  think 
of  enzymes  as  being  amphoteric  substances,  in  some  of  which  the 
positive  charge  predominates,  in  others,  the  negative;  as  a  corollary 
of  this,  pepsin  dissolves  in  alkaline  solution;  in  other  words,  the  pepsin 
is  dissolved  away  from  the  substrate  it  would  digest  and  is  thus  in- 
activated; the  reverse  of  this  holds  true  for  trypsin.  Salivary  dia- 
stase seems  entirely  neutral,  since  saliva  must  fluidify  as  readily  in 
acid  as  in  alkaline  reaction.  In  some  cases  we  observe  a  relationship 
between  the  reaction  of  the  substrate  upon  which  a  ferment  is  to 
act  and  the  ferment;  thus,  pepsin-rennin  has  a  pronounced  basic 
character,  casein  an  acid  character,  albumin  in  acid  solution  has  a 
basic  character,  and  as  such  combines  with  acid  pepsin;  in  alkaline 
solution  it  has  an  acid  character  and  so  can  unite  with  basic  trypsin. 
Consequently,  we  find  here  phenomena  which  I  pointed  out  in  my 
experiments  on  the  adsorption  of  dyestuffs,  page  29. 

This  difference  in  adsorbability  is  utilized  in  many  cases  for  the 
'purification  of  enzymes.  Thus  L.  Michaelis  removed  albumin  from 
mixtures  of  serum  albumin  and  invertin  by  shaking  them  in  acid 
solution  with  kaolin;  the  invertin  remained  without  loss  of  strength 
in  the  albumin-free  solution.  E.  Abderhalden  and  F.  W.  Strauch 
extracted  pepsin  from  the  stomach  content  of  animals  by  means  of 
elastin  and  then  recovered  it  from  the  elastin  by  means  of  water. 

Depending  upon  the  reaction,  decided  differences  were  found 
when  enzymes  were  filtered  through  Chamberland  filters.  Accord- 
ing to  Holderer  most  of  those  studied  by  him  passed  through  the 
filter  when  they  were  neutral  to  Phenolphthalein  but  they  were  held 
back  when  neutral  to  methyl  orange.  Holderer  attributes  this 
principally  to  the  effect  of  adsorption  by  the  filter  mass. 

For  a  number  of  enzymes  the  course  of  the  reactions  was  studied 
and  proved  to  be  quite  complicated.  I  refer  to  the  investigations  of 
platinum  sol  by  G.  Bredig  and  his  pupils;  of  invertase  and  amylase 
by  V.  Henri;  of  lipase  by  M.  Bodenstein  and  Dietz  and  of  emulsin 
by  P.  Jacobson,  which  are  described  by  H.  Freundlich  in  his 
"  Kapillar chemie."  It  is  conclusively  shown  in  the  publication  of 
W.  S.  Denham*  from  Bredig's  Institute  that  among  all  the  compli- 
cated factors,  it  is  the  surface  concentration  which  is  of  the  greatest 
importance  for  the  acceleration  of  the  reaction. 

In  the  case  of  gels,  the  greater  the  surface  concentration  becomes,  or 
in  other  words,  the  more  swollen  the  substrate  is,  the  better  is  the 
opportunity  offered  an  enzyme  to  enter  the  substrate.  This  obser- 
vation can  be  made  over  and  over  again  in  the  cases  of  enzyme 
cleavage.     E.  Knoevenagel*  offers  convincing  proof  of  this  fact. 


188  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

He  says  "The  degree  to  which  acetyl  cellulose  swells,  runs  parallel 
with  its  speed  of  saponification  by  aqueous  alkalis,  so  that  with 
greatly  swollen  acetyl  celluloses  the  saponification  by  1/2  n  KOH  is 
completed  quantitatively  in  a  few  hours,  at  room  temperature." 

We  have  not  yet  answered  the  question :  What  property  is  to  be 
ascribed  to  the  specific  action  of  enzymes?  We  may  regard  this 
query  as  being  answered  by  G.  Bredig  and  Fajans*  as  far  as  the 
principle  involved  is  concerned.  They  demonstrated  that  right 
and  left  campho-  and  bromo-campho-carbonic  acid  (which  resemble 
one  another  like  a  picture  and  its  reflection  in  a  mirror)  may  be 
split  into  camphor  and  carbonic  acid  by  bases  acting  as  catalyzers. 
The  speed  with  which  these  two  opposites  are  broken  down  differs 
considerably  if  optically  active  bases  (quinine,  quinidine,  nicotine 
and  cinchonine)  are  employed,  and  may  amount  to  50  per  cent. 
This  is  analogous  to  the  specific  action  of  enzymes  upon  chemically 
known  substances  with  catalyzers  which  have  definite  chemical 
characteristics.  It  has  been  shown  for  most  enzyme  actions,  with 
certain  exceptions  among  the  sugars,  that  of  two  optically  active 
isomers  both  are  attacked  by  a  given  enzyme,  but  one  is  always 
affected  more  quickly.  We  thus  have  a  complete  analogy  for  natural 
enzymes,  but  beyond  this  point  the  "lock  and  key"  idea  fails,  since 
under  no  circumstances  could  "an  asymmetric  key  fit  the  mirror 
image  of  its  proper  lock." 

But  the  analogy  extends  further.  G.  Bredig  and  Fiske  effected 
asymmetric  synthesis  by  a  catalyzer  of  known  composition.  According 
to  L.  RosE-NTHALER,  the  enzyme  action  of  emulsin  accelerates  the 
following  reaction: 

C6H5.CHO+    HON    =  CeHs  •  CH(OH)  •  ON. 

Benzaldehyde      +  hydrocyanic,  nitromandelic  acid, 

acid 

G.  Bredig.  and  Fiske  replaced  emulsin  with  quinme,  but  if  they 
employed  Chinidin  as  a  catalyzer  they  obtained  laevo  rotary  nitro- 
mandeHc  acid  in  addition  to  the  inactive  product. 

We  may  summarize  our  present  understanding  of  enzyme  action 
thus:  As  a  result  of  their  colloidal  properties,  under  favorable  ex- 
ternal circumstances,  enzyme  and  substrate  are  greatly  concentrated 
at  their  interfaces,  so  that  the  course  of  the  reaction  is  very  much 
accelerated;  the  reaction  between  enzyme  and  substrate  is  purely 
chemical,  conditioned  by  their  mutual  chemical  constitution  or  con- 
figuration. [Ultramicroscopic  observations  suggest  that  possibly 
physical  action  is  also  involved.  J.  Alexander,  Jour.  Am.  Chem. 
Soc,  1910,  vol.  32,  p.  680.     Tr.] 

Enzymes,  perhaps,  exhibit  the  property  of  aging  to  a  greater  ex- 
tent than  all  other  colloids.     Some,  e.g.,  trypsin,  if  dried,  lose  their 


ENZYMES  189 

activity  after  a  time;  in  solution  they  all  deteriorate  more  or  less 
rapidly.  We  do  not  know  whether  this  depends  upon  purely  me- 
chanical variations,  or  whether  it  is  associated  with  a  chemical 
change.  For  the  former  view  we  have  the  fact  that  many  enzyme 
solutions  may  be  inactivated  by  mere  shaking;  a  rennet  solution,  for 
instance,  need  be  violently  shaken  only  two  minutes  in  a  test  tube 
in  order  largely  to  deprive  it  of  its  capacity  to  coagulate  milk.  Even 
E.  Abderhalden  and  M.  Guggenheim*  had  observed  that  tyro- 
sinase, expressed  yeast  juice,  and  pancreatic  juice  had  their  activity 
partially  inhibited  by  shaking  them  for  24  hours.  A.  O.  Shaklee 
and  S.  J.  Meltzer*  found  the  same  true  for  pepsin  and  M.  M. 
Harlow  and  P.  G.  Stiles*  for  ptyalin. 

Quite  independently  in  1908,  Signe  and  Sigval  Schmidt-Nielsen* 
observed  the  inadivation  of  rennet  by  shaking,  and  subjected  the 
phenomenon  to  a  thorough  study.  It  was  deduced  from  this  that 
inactivation  by  shaking  is  a  surface  phenomenon;  the  inactivation  in- 
creases with  the  length  of  time  and  the  violence  of  the  shaking;  the 
volume  of  air  present,  the  concentration  of  the  enzyme,  and  the  tem- 
perature are  all  influencing  factors.  The  enzyme  becomes  con- 
centrated in  the  foam  and  on  the  surface  of  the  vessel  employed. 
The  foam  is  more  active  than  the  fluid  and  the  procedure  offers  a 
possible  method  of  concentrating  enzymes.  If  a  rennet  solution  that 
has  been  shaken  is  allowed  to  stand,  it  recovers  some,  but  never  all 
of  its  original  activity;  a  portion  remains  irreversible.  If  saponin 
is  added  to  a  rennet  solution,  no  inactivation  results  from  shaking 
because  saponin  drives  the  rennet  from  the  surface. 

Subsequently  M.  Jacoby  and  A.  Schütze*  published  an  analogous 
observation.  They  found  that  hemolytic  complement  (see  p.  196)  of 
guinea-pig  serum  was  inactivated  by  shaking  it  at  37°  C.  Reac- 
tivation, in  other  words  the  reversibility  of  the  process,  depends 
on  the  duration  of  the  shaking.  At  first,  only  a  definite  fraction 
of  the  complement  is  irreversibly  inactivated  by  the  shaking  since 
it  may  be  reactivated  by  "end  piece"  and  also  incompletely  by 
"middle  piece."  When  the  shaking  has  been  sufficiently  prolonged 
the  complement  is  irreversibly  inactivated  according  to  Ritz.  The 
inactivation  depends,  according  to  P.  Schmidt  and  Lieber,  on 
the  fact  that  the  serum  is  made  turbid  by  shaking,  a  foam  is 
formed  into  which  the  globulin  separates,  and  this  globuHn  adsorbs 
the  complement.  The  reactivation  by  "end  piece"  results  from  the 
solution  of  the  flocculated  globulin  thus  liberating  the  complement 
(see  p.  196).  To  what  extent  the  action  of  the  alkali  (from  the  glass) 
assists  in  the  inactivation  has  not  been  determined  with  certainty. 
On  the  contrary,  it  seems  from  the  data,  that  only  a  portion  of  the 


190  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

complement  is  inactivated,  since  it  may  be  reactivated  by  addition 
of  ''end  piece"  and  of  ''middle  piece."  Serum  becomes  turbid  on 
shaking,  and  it  is  the  author's  opinion  that  this  is  evidently  due  to  a 
coagulation  of  the  albumin  by  shaking. 

In  many  investigations,  especially  in  "immunity  studies,"  it  is  cus- 
tomary to  shake  the  test  tubes,  and  I  believe  that  some  of  the  dis- 
agreements in  the  results  of  experiments  by  different  investigators 
are  due  to  a  disregard  of  such  surface  phenomena.  [R.  Ottenberg 
does  not  consider  this  factor  in  his  exhaustive  study.  Arch,  of  Int. 
Medicine,  vol.  xix,  pp.  457-492.     Tr.] 

The  diffusion  coefficient  of  several  enzymes  was  measured  ^  by  R.  0. 
Herzog  and  H.  Kasarnowski.*     They  are  for 

Pepsin 0.062  (at  12°  C.) 

Pepsin 0.066  (at  16°  C.) 

Rennin 0.062  (at  16°  C.) 

Invertin 0.032  (at  16*  C.) 

Emulsin 0.033  (at  15.3°  C.) 

From  these  figures  the  following  molecular  weights  were  cal- 
culated : 

Pepsin 13,000 

Invertin 54,000 

Emulsin 45,000  ] 

In  studies  on  the  filtration,  ultrafiltration  and  diffusion  of  enzymes 
through  membranes  it  must  be  determined  beforehand,  whether  the 
filter  adsorbs  too  strongly.  Thus,  e.g.,  a  Chamberland  filter  per- 
mits no  pepsin,  trypsin,  lipase  or  zymase  to  pass  through,  though 
the  pores  are  of  ample  size.  By  choosing  suitable  membranes, 
these  methods  of  separation  have  given  valuable  results.  It  has 
been  possible  by  diffusion  and  ultrafiltration  to  separate  a  num- 
ber of  enzymes,  which  were  formerly  regarded  as  individual,  into 
two  constituents  having  different  properties.  Thus,  according  to 
S.  Fraenkel  and  M.  Hamburg,*  diastase  prepared  from  malt  may 
be  divided  into  two  enzymes.  The  one  which  diffuses  changes 
starch  into  sugar,  whereas  the  other  merely  fluidifies  the  starch. 
A.  VON  Lebedew*  ultrafiltered  expressed  yeast  juice  and  thus 
succeeded  in  demonstrating,  that  in  fermenting  sugar  the  disap- 
pearance of  the  sugar  and  the  formation  of  carbonic  acid  are  two 
distinct  processes. 

In  the  course  of  such  experiments  it  has  been  shown  frequently, 
that  the  components  are  inactive  individually  and  only  exert  their 
enzyme  action  in  combination.  The  first  observation  of  this  kind 
was  that  of  R.  Magnus,*  who  dialyzed  liver  extract.  The  extract 
which  originally  split  up  fat  thus  became  inactive;  when  Magnus 
*  The  figures  are  the  mean  of  several  determinations. 


ENZYMES  191 

J- 
united  residue  and  dialyzate,  the  mixture  recovered  its  lypolytic 
properties.  A.  Harden  and  W.  J.  Young*  made  a  similar  observa- 
tion when  they  ultrafiltered  expressed  yeast  juice.  The  filter  res- 
idue lost  its  ability  to  cause  fermentation  but  regained  it  when 
mixed  with  the  filtrate.  It  is  evident  from  this,  that  some  enzymes 
consist  of  a  colloid  and  a  crystalloid  constituent;  the  latter  follow- 
ing the  suggestion  of  G.  Bertrand  is  called  co-enzyme  or  co-ferment. 
In  still  other  ways  the  co-enzyme  shows  crystalloid  properties; 
unlike  the  colloid  portion  it  is  insensitive  to  boiling  and  frequently 
consists  of  a  substance  whose  composition  is  well  known.  Thus, 
e.g.,  according  to  0.  von  Fürth  and  J.  Schütz,*  sodium  cholate  and 
sodium  glycocholate  are  co-enzymes  of  lipase;  and  according  to 
BiERRY  and  V.  Henri*  the  chlorin  and  bromin  ions  of  alkaline  salts 
are  the  co-enzymes  for  the  action  of  pancreatic  juice  upon  starches. 

In  contradistinction  to  the  co-enzymes,  the  anti-enzymes  are 
usually  colloids.  Anti-enzymes  are  substances  which  interfere  with 
the  action  of  enzjmies.  They  are  like  the  antitoxins  which  detoxi- 
cate  toxins,  and  like  the  antitoxins,  they  occur  to  some  extent  in 
normal  serum,  or  may  be  produced  in  it  by  the  injection  of  enzymes. 
For  instance,  horse  serum  contains  a  large  amount  of  anti-rennin 
which  inhibits  the  coagulation  of  milk  by  rennin.  By  injecting  the 
proper  enzyme,  anti-enzymes  for  lipase,  emulsin,  amylase,  pepsin, 
papain  and  urease  have  been  obtained.  An  exception  to  this  is  anti- 
trypsin which  seems  to  be  a  crystalloid  since  it  diffuses  readily.  It 
is  the  anti-enzyme  which  protects  intestinal  parasites  from  digestion 
by  the  pancreatic  juices. 

According  to  S.  G.  Hedin,*^  the  relationship  between  enzjrme  and 
anti-enzyme  is  an  adsorption,  which  probably  results  in  a  fixation. 

A  certain  similarity  between  enzyme  and  co-enzjntne  is  possessed 
by  pro-enzyme  and  its  activator.  Most  enzymes  are  formed  in  an 
inactive  state  called  the  pro-enzyme,  pro-ferment,  or  zymogen,  which 
becomes  active  only  after  the  addition  of  some  crystalloid,  usually  a 
simple  substance.  The  pro-enzyme  of  pepsin  may  be  extracted  from 
the  gastric  mucous  membrane,  but  cannot,  digest  albumin;  only  after 
the  addition  of  very  dilute  acids  does  it  become  pepsin  and  acquire 
its  ability  to  digest.  The  trypsin  of  pancreatic  juice  is  excreted  into 
the  duodenum  as  an  inactive  pro-enzyme;  it  is  activated  by  calcium 
salts.  This  is  of  the  greatest  biological  significance,  since  otherwise, 
secreting  glands  would  not  be  safe  from  their  own  secretions. 

According  to  E.  Pribram,*  the  formation  of  pro-enzymes  occurs 
in  this  manner;  the  protoplasm  of  the  glandular  cells  retains  a  cer- 
tain portion  of  the  food  by  adsorption.  Acids,  calcium  salts,  etc., 
arrest  the  adsorption,  and  the  active  ferment  becomes  free.    In 


192  COLLOIDS  IN   BIOLOGY  AND  MEDICINE 

support  of  this  view,  E.  Pribram  and  E.  Stein  injected  through  a 
tube  into  the  stomach  of  one  rabbit  an  active  solution  of  rennin, 
and  into  the  other,  one  mactivated  by  boihng.  After  four  hours  the 
rabbits  were  killed  and  the  amounts  of  pro-ferment  contained  in  their 
gastric  mucous  membranes  were  measured.  The  gastric  mucous  mem- 
brane of  the  rabbit  treated  with  active  rennin  contained  much  more 
pro-enzyme  than  the  other.  From  this,  the  authors  deduced  that  the 
colloidal  gastric  mucous  membrane  adsorbed  the  rennin  and  changed 
it  into  pro-enzyme.  S.  G.  Hedin*  also  views  zymogen  as  the  com- 
bination of  an  enzyme  (rennet)  with  an  inhibiting  substance.  In  the 
condition  studied  especially  by  him,  rennet  is  freed  by  hydrochloric 
acid,  the  inhibiting  substance  is  let  free  by  ammonia,  with  the 
destruction  of  the  rennet. 


CHAPTER  XIII. 
IMMUNITY  REACTIONS. 

That  the  organism  is  overwhelmed  by  a  large  dose  of  poison  but 
recovers  from  a  small  one  should  not  particularly  surprise  us.  Ever 
smce  the  recognition  of  the  nature  of  infectious  diseases,  it  must  have 
amazed  biologists  that  every  infected  organism  did  not  succumb  to 
the  sUghtest  infection.  Microorganisms  multiply  indefinitely,  and, 
theoretically,  it  is  only  a  question  of  hours  before  the  number  present 
shall  be  overwhelming  whether  the  infection  is  with  a  large  or  a  small 
dose.  Were  this  assumption,  to  which  we  might  be  led  from  the  ob- 
servation of  culture  media,  correct,  no  living  thing,  plant  or  animal, 
could  exist.  There  must  be  inherent  forces  in  the  living  organism 
which  protect  it  agaihst  pathogenic  germs,  which  make  it  immune  to 
such  injuries,  and  which  are  called,  accordingly,  immune  bodies 
(immune  substances). 

L.  Pasteur  was  the  pioneer  in  the  systematic  study  of  immunity. 
He  produced  experimental  proof  that  immunity  might  be  artificially 
produced  by  previous  treatment  with  attenuated  infective  agents 
(chicken  cholera)  just  as  had  been  done  previously,  in  the  case  of 
vaccination  against  smallpox.  These  investigations  received  a 
mighty  impulse  when  Robert  Koch  succeeded  in  growing  disease 
germs  in  pure  culture.  The  doctrines  of  immunity  and  predisposition 
were  developed  into  a  special  branch  of  science  which  at  present 
holds  the  chief  interest  of  scientific  medicine. 

It  was  recognized  that  the  body  could  overcome  its  invaders  in 
various  ways:  substances  occur  which  make  bacteria  harmless  by 
dissolving  them,  bacteriolysins  (acting  against  vibrios,  e.g.,  of  cholera, 
and  against  typhoid),  and  others  which  clump  them  together  and 
precipitate  them,  the  agglutinins  (against  typhoid,  paratjq^hoid, 
dysentery,  etc.).  In  other  cases,  the  protection  is  directed  prin- 
cipally against  the  poisons,  toxins,  which  the  organized  germs  de- 
velop (diphtheria  toxin,  tetanotoxin,  etc.).  The  organism  possesses 
a  peculiar  protective  mechanism  in  the  leucocytes,  which  take  up 
and  digest  the  bacteria  and  cocci,  devouring  them  like  free  living 
amebse  in  search  of  food.  This  phenomenon,  which  was  recognized 
and   studied   chiefly  by  E.  Metschnikoff,  is  called  phagocytosis. 

193 


194  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  bacteria  are  previously  prepared  for  phagocytosis  by  certain 
Immune  substances  in  the  serum  called  opsonins. 

The  study  of  these  phenomena  was  very  much  simplified,  when  it 
became  possible  to  transfer  many  of  them  from  the  living  organism 
to  test  tubes.  They  were  thus  freed  from  disturbing  epiphenomena 
and  made  susceptible  to  quantitative  investigations.  By  these 
methods  of  study,  we  have  learned  a  number  of  properties  of  the 
blood  and  cells,  which  have  no  direct  influence  on  the  natural  pro- 
tection of  the  organism  against  the  attacks  of  microorganisms,  or 
which  may  be  regarded  merely  as  epiphenomena.  They  lead  to  the 
knowledge  that  the  weapons  of  the  organism  against  disease  germs 
are  not  teleologically  forged  for  this  sole  purpose,  but  that  they  are 
the  product  of  a  universal  biologic  law,  according  to  which  the 
organism  produces  antisubstances  against  all  kinds  of  substances 
foreign  to  the  species  (art-fremde). 

In  accordance  with  their  historic  recognition,  and  the  method  of 
their  investigation,  it  is  customary  to  class  them  with  immunity  phe- 
nomena: I  am  referring  to  the  substances  which  dissolve  and  floccu- 
late blood  corpuscles,  the  hemolysins  and  hemagglutinins  and  the 
albumin-precipitating  substances,  the  precipitins:  and  finally  the 
Wassermann  reaction  in  syphilis,  and  anaphylaxis.    . 

If  the  sera  of  two  animals,  e.g.,  cattle  serum  and  rabbit  serum,  are 
mixed,  the  solution  remains  clear.  If  an  animal,  e.g.,  a  rabbit,  is  in- 
jected with  the  serum  from  a  different  species  of  animal,  e.g.,  cattle 
serum,  substances  are  formed  in  the  rabbit,  precipitins.'^  If  we  then 
mix  the  serum  of  such  an  animal,  ''cattle-rabbit,"  with  ox  serum,  a 
precipitate  forms.  Agglutinins  and  hemolysins  develop  in  a  similar 
way.  If  a  rabbit  has  cattle  blood  corpuscles  injected  into  its  veins, 
substances  develop  in  the  rabbit  serum  which  agglutinate  and  dissolve 
the  cattle  blood  corpuscles.  Hemolysin  consists  of  two  substances, 
one  heat  resisting  (thermostable)  and  specific,  the  amboceptor,  and 
another,  heat  sensitive  (thermolabile,  destroyed  at  55°  C.)  and  non- 
specific, the  complement.  Only  the  amboceptor  develops  as  a  result 
of  injecting  the  red  blood  corpuscles,  the  complement  is  always 
present  in  every  serum.  However,  both  are  required  for  hemolysis. 
We  have  now  explained  the  formation  of  precipitins  for  cattle  serum 
or  blood  corpuscles  in  rabbits,  but  the  principle  is  of  general  applica- 
tion for  the  injection  of  blood  into  a  different  species  of  animals.     To 

^  The  so-called  "precipitin  reaction"  is  of  great  medico-legal  importance. 
It  serves  for  the  differentiation  oi  human  and  animal  blood,  for  which  a  small 
drop  suffices.  It  is  also  employed  in  detecting  adulterations  (horse-meat  in 
sausage,  etc.).  In  the  study  of  phylogenesis  it  is  a  valuable  aid  particularly  in 
teaching  the  natural  relationships  of  animals. 


IMMUNITY  REACTIONS  195 

make  the  application  general,  if  an  animal  is  injected  with  sub- 
stances foreign  to  its  species  {antigen),  e.g.,  albumin,  animal  cells, 
bacteria,  toxin,  antibodies  or  immune  substances  (precipitins,  hemol- 
ysins, agglutinins,  antitoxins)  are  formed  in  the  injected  animal. 

Binding  of  antigens  (toxin,  bacteria,  etc.)  by  the  immune  sub- 
stances (antitoxin,  bacteriolysin,  etc.)  results  from  combination  or  a 
sort  of  neutralization  which  may  be  compared  to  the  neutralization 
of  an  acid  by  a  base.  P.  Ehrlich*  was  the  first  to  study  this  neu- 
tralization quantitatively  and  showed  in  the  case  of  diphtheria  toxin 
and  its  antitoxin,  that  the  saturation  did  not  occur  as  in  the  case  of 
a  strong  acid,  e.g.,  HCl,  and  a  strong  base,  e.g.,  KOH,  but  that  the 
diphtheria  toxin  must  consist  of  a  mixture  of  more  or  less  acid  toxins. 
We  reach  this  conclusion,  not  only  from  the  course  of  the  saturation 
curve,  but  also  from  a  study  of  the  different  poisonous  actions  pos- 
sessed by  the  individual  saturation  fractions.  Though,  e.g.,  the  larg- 
est part  of  the  diphtheria  toxin  has  an  acute  toxic  action,  there  is  a 
particular  fraction,  the  toxon,  which,  after  two  or  three  weeks,  pro- 
duces paralysis  of  the  extremities  that  are  quite  foreign  to  the  toxin. 
In  different  cases  various  indicators  are  used  as  a  sign  of  the  union 
between  antigen  and  immune  substances.  In  the  case  of  toxin-antitoxin  ^ 
we  are  reduced  to  the  biological  proof  by  animal  experiments;  the  re- 
duction in  the  toxicity  of  the  mixtures  is  determined  from  the  toxic 
action  remaining  in  them.  In  the  case  of  hemolysins,  the  ability  to 
dissolve  red  blood  cells  more  or  less  completely  is  used  as  a  sign. 
Precipitins  are  recognized  by  testing  the  antigen  against  various 
dilutions  and  determining  the  greatest  dilution  at  which  turbidity 
can  still  be  recognized.  If,  e.g.,  a  ra;bbit  has  been  injected  with 
goat  serumi  a  substance  develops  in  the  rabbit  which  precipi- 
tates goat  serum.  If  we  add  to  goat-rabbit  serum  which  has  been 
placed  in  a  row  of  test  tubes  goat  serum  in  a  dilution  1/100, 
1/1000,  1/10000,  etc.,  we  shall  find  a  dilution  at  which  merely  tur- 
bidity occurs.  In  a  similar  manner  agglutinins  are  tested  (in  this 
instance  the  immune  serum  is  diluted). 

Since  the  nomenclature  is  not  uniform,  a  table  of  the  terms  in  com- 
mon use  is  given  here. 

Agglutinins  change  bacteria  so  that  they  may  even  be  precipitated  by  alkali 
salts  (see  antigen). 

Amboceptor,  see  hemolysin. 

Antigens,  foreign  substances  (bacteria,  proteins,  toxins,  etc.)  against  which 
specific  antisubstances  (antibodies)  are  developed  by  an  animal  injected  with 
them  (agglutinins,  precipitins,  antitoxins,  etc.). 

Antibodies,  immune  bodies. 

Antitoxin,  specific  antibodies  which  neutrahze  toxins. 

End  piece,  see  hemolysin. 

^  [Jerome  Alexander  observed  in  the  ultramicroscope  the  mutual  coagulation 
of  diphtheria  toxin  and  antitoxin  and  tetanus  toxin  and  antitoxin;  diphtheria 
toxin  was  not  precipitated  by  tetanus  antitoxin.     Tr.] 


196  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Hemolysins  dissolve  red  blood  corpuscles.  Two  substances  are  usually  required 
for  hemolysis.  One  is  specific,  the  real  antibody,  and  is  called  amboceptor.  The 
other  occurs  in  every  serum  and  is  complement.  Complement  consists  of  two 
parts,  one  of  which,  the  middle  piece,  is  precipitated  with  the  globulin;  the  end 
piece  remains  with  the  albumin  of  the  serum.  Only  when  both  are  united  does 
complement  act.  According  to  H.  Sachs,  Omokvkow  and  Ritz  there  is  an  addi- 
tional "  third  component,"  quite  heat  stable.  According  to  P.  Schmidt  comple- 
ment is  a  single  substance  of  which  a  portion  is  adsorbed  by  globuhn  when  it 
is  precipitated. 

Complement,  see  hemolysin. 

Lysins  cause  solution.  Bacteriolysins  dissolve  bacteria,  hemolysins  dissolve 
red  blood  corpuscles. 

Precipitins  flocculate  albumin. 

Toxins,  poisons  which  produce  antitoxins  when  injected. 

The  Nature  of  Antigens  and  Immune  Bodies. 

The  substances  involved  in  immunity  reactions  are  all  dissolved 
or  suspended  colloids.  There  is,  therefore,  a  particular  reason  for 
studying  these  questions  from  the  standpoint  of  colloid  investigation.^ 

So  far  it  has  been  impossible  to  produce  immune  bodies  by  means 
of  a  crystalloid;  a  foreign  colloid  (antigen)  has  always  been  required. 

The  proof  of  the  colloid  character  of  antigens  and  immune  bodies 
has  been  demonstrated  in  numerous  cases.  Upon  dialysis,  they 
do  not  pass  through  a  dialyzing  membrane;  the  diffusibility  of  diph- 
theria toxin  and  tetanolysin  and  their  antitoxin  are  indicative  of  a 
particle  magnitude  of  the  same  order  as  hemoglobin  (Sv.  Arrhenius). 
Ultrafiltration  of  diphtheria  toxin,  toxon  and  antitoxin  and  anti-rennin 
gave  similar  results  (H,  Bechhold).  The  hemolytic  complement  of 
guinea-pig  serum  is  inactivated  by  shaking  with  the  formation  of  a 
precipitate  (M.  Jacob y  and  A.  Schütze).  This  indicates  a  concen- 
tration at  the  boundary  of  fluid/air,  as  in  the  case  of  albumin  and 
other  colloids.  The  observation  of  W.  Biltz,  H.  Much  and  C. 
SiEBERT  that  a  bactericidal  horse  serum  loses  its  bactericidal  activity 
upon  shaking  is  to  be  ascribed  to  a  similar  phenomenon.^ 

^  We  may  mention  the  following  papers  which  treat  Immunity  with  particu- 
lar reference  to  the  standpoint  of  coUoid  chemistry: 

K.  Landsteiner,  Die  Theorien  der  Antikörperbildung,  Wiener  Klin.  Wochen- 
schr.  22,  Nr.  47   (1909). 

Idem.,  Kolloide  u.  Lipoide  in  d.  Immunitätslehre  im  Handbuch  d.  Pathogenen 
microorganism  von  Kolle  u.  Wassermann,  Bd.  II  (1913). 

O.  PoRGES,  im  Handb.  d.  Technik  u.  Methodik  d.  Immunitätsforschimg,  Bd.  II, 
Lief.  2  (Jena,  1909). 

H.  Zangger,  Viertel] ahrsschr.  d.  Naturf.-Ges.  in  Zurich,  1908,  408-455. 

2  It  might  be  claimed  that  these  substances,  which  it  is  impossible  to  prepare  in 
pure  form,  are  not  in  themselves  colloids,  but  that  they  are  adsorbed  by  the 
proteins  simultaneously  present  in  the  solution  and  thus  simulate  -what  coUoid 
character  they  exhibit.  For  the  correctness  of  this  view  no  evidence  tas  hitherto 
been  presented. 


IMMUNITY  REACTIONS  197 

It  is  held  by  one  small  group  of  investigators  that  antigens  are 
hpoids  or  lipoid-albumin  compounds.  Since  the  part  taken  by  lipoids 
in  many  immunity  reactions  is  not  definitely  settled,  it  is  impossible 
as  yet  to  determine  the  general  correctness  of  this  view.  At  any 
rate  it  has  not  as  yet  been  possible  to  immunize  with  the  lipoids 
chemically  known. 

Since  our  knowledge  concerning  the  chemical  composition  of 
normal  proteins  is  still  meager,  what  we  know  about  the  proteins  of 
immune  bodies  cannot  be  more  ample.  According  to  Kirschbaum 
dysentery  toxin  is  acid.  By  ultrafiltration,  he  prepared  water  insolu- 
ble acid  dysentery-toxin  which  was  nearly  atoxic,  though  the  salt 
obtained  by  dissolving  the  acid  in  an  alkaline  carbonate  possessed 
the  poisonous  properties  of  the  toxin.  The  experiments  of  Fr. 
Obermayer  and  E.  P.  Pick*  indicate  that  the  aromatic  nucleus  in 
the  antigen  is  of  great  importance  for  the  development  and  character 
of  the  antibodies. 

Antibodies  are  universally  regarded  as  albuminous. 

Before  we  discuss  details,  let  us  indicate  a  great  misunderstanding 
which  at  the  time  gave  rise  to  heated  discussions  and  clouded  the 
issues.  The  quantitative  relations  in  which  the  substance  in  ques- 
tion enters  into  reaction  (toxins  with  their  antitoxins,  bacteria  with 
their  agglutinins,  etc.)  have  great  similarity  to  adsorption  curves 
(W.  BiLTz)  and  to  the  neutralization  curves  of  certain  weak  acids 
and  bases  (Sv.  Arrhenius).  These  investigators  laid  great  stress 
on  this  fact  and  believed  that  they  had  thus  discovered  the  nature 
and  the  course  of  the  immunity  reactions  in  question.  P.  Ehrlich 
raised  the  weighty  objection,  that  the  reaction  is  specific,  and  that 
the  poisons  are  very  complex:  diphtheria  toxin  is  detoxicated  only 
by  diphtheria  antitoxin;  typhoid  bacilli  are  precipitated  only  by 
typhoid  agglutinin.  There  is  no  doubt  that  these  specific  proc- 
esses cannot  be  explained  by  what  we  call  colloid-chemical  reactions 
(see  H.  Bechhold*^)  .  We  must  conceive  of  the  process  as  occurring 
in  two  stages,  and  we  must  emphasize  that  this  sharp  distinction 
does  not  obtain  in  every  case. 

First  Stage:  The  two  colloids,  toxin  and  antitoxin,  bacterium  and 
agglutinin,  unite  in  accordance  with  the  laws  governing  other  colloids, 
e.g.,  fiber  and  dye,  and  the  specific  substances  react  on  one  another 
and  it  is  still  an  open  question  whether  we  must  represent  these 
reactions  as  chemical  or  catalytic. 

Second  Stage.  The  colloidal  product  of  the  reaction  shows  physical 
properties  which  distinguishes  it  from  the  reacting  substances,  e.g., 
it  precipitates. 

We  cannot  enter  here  into  the  question  of  specific  combination. 


198  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

A.   The  Distribution  of  Immune  Substances  Between 
Suspensions  and  Solvent. 

Bacteria  and  blood  corpuscles  form  suspensions  which  to  a  greater 
or  less  extent  are  able  to  attract  to  themselves  immune  substances. 
It  is  fortunate  for  the  study  of  these  phenomena  that  many  experi- 
ments have  been  preformed  upon  the  adsorption  of  inorganic  sus- 
pensions (kaolin,  charcoal,  ferric  hydroxid  gel,  etc.)  from  solutions 
of  known  composition.  For  comparison  many  investigators  have  per- 
formed appropriate  experiments  on  toxins  and  immune  substances. 

Adsorption  by  Means  of  Inorganic  Suspensions  and  Hydrogels. 
A  sign  of  adsorption  is  a  strong  withdrawal  of  a  dissolved  substance 
from  dilute  solutions  and  a  relatively  smaller  withdrawal  from  such 
as  are  more  concentrated.  This  requires  extensive  quantitative  inves- 
tigation with  solutions  of  different  concentration.  Unfortunately, 
there  are  but  few  such  experiments  published.  It  may,  however,  be 
deduced  from  the  results  of  some  of  these  experiments  that  an  ad- 
sorption curve  is  actually  involved. 

Even  W.  Roux  and  Yersin*  found  that  calcium  phosphate, 
aluminum  hydroxid  and  bone  black  removed  some  poison  from  a 
solution  of  diphtheria  toxin,  but  that  the  solution  was  never  entirely 
detoxicated.  W.  Biltz,  H.  Much  and  C.  Siebert*  shook  gels  of 
iron  oxid,  chromium  oxid  and  zirconium  oxid,  among  others,  ^s^ith 
tetanus  and  diphtheria  toxin,  tetanolysin  and  a  bactericidal  horse 
serum.  They  determined  a  diminution  in  the  activity  of  the  respec- 
tive solutions,  and  that  for  the  same  quantity  of  hydrogel,  the 
diminution  by  activity  was  frequently  more  marked  for  dilute  than 
for  concentrated  solutions.  Occasionally  complete  fixation  or  destruc- 
tion occurred,  e.g.,  in  the  case  of  typhoid  agglutinin.  H.  Bechhold** 
found  that  arachnolysin  and  staphylolysin  were  never  completely  re- 
moved from  solution  by  formol-gelatin  or  cellulose.  K.  Land- 
STEiNER  and  his  pupils  shook  tetanus  toxin  with  kaolin,  protagon, 
Cholesterin,  palmitic  acid,  stearic  acid  and  lecithin;  a  poisonous  re- 
siduum was  always  discovered  in  the  solution. 

Since  complement  may  be  removed  from  a  solution  by  various  sus- 
pensions (for  literature  see  H.  Sachs),  a  mechanical  adsorption  is 
probable. 

Specific  Adsorption. 

Glancing  at  the  entire  literature  on  this  question,  we  are  con- 
fronted with  the  great  difference  in  adsorption  capacity  of  the  ad- 
sorbents as  well  as  of  the  adsorbed  substances.  Although  tetanus 
toxin  is  well  adsorbed  by  kaolin,  protagon,  Cholesterin,  palmitic  acid, 


IMMUNITY  REACTIONS  199 

stearic  acid,  and  lecithin,  only  very  little  of  it  is  taken  up  by  cetyl 
alcohol,  casein,  coagulated  serum  albumin,  and  starches  (K.  Land- 
STEiNER  and  A.  Botteri*)  .  Arachnolysin  is  adsorbed  more  strongly  by 
glacial  acetic  acid  collodion  than  by  formol-gelatin;  glacial  acetic  acid 
collodion  adsorbs  rennin  very  strongly,  but  adsorbs  practically  none 
of  a  serum  containing  anti-rennin  (H.  Bechhold*^).  Silicic  acid  and 
barium  sulphate  fix  complement  which,  however,  is  also  fixed  by 
kaolin  to  a  lesser  extent  (E.  Hauler*). 

In  view  of  these  specific  influences,  L.  Jacque  and  E.  Zunz*  un- 
dertook extensive  experiments  upon  the  adsorption  of  antigens  and 
antibodies  by  inorganic  suspensions.  They  concluded,  as  had  pre- 
viously been  shown  by  E.  Zunz*  that  differences  in  surface  tension 
were  not  alone  determinative  for  adsorption.  They  found,  e.g.,  that 
bone  black  strongly  adsorbed  diphtheria  toxin  as  well  as  antitoxin, 
though  neither  was  adsorbed  by  wood  charcoal,  diatomaceous  earth, 
talc,  kaolin  or  clay.  Nevertheless  kaolin  and  clay  adsorb  tetanolysin. 
Bone  black,  a  good  adsorbent  for  diphtheria  antitoxin,  does  not 
adsorb  the  antitoxin  of  tetanolysin  or  cobra  hemolysin. 

Reversibility.  A  purely  mechanical  adsorption  demands  that  the 
process  be  completely  reversible.  This  occurs  in  the  case  of  the 
slightest  adsorptions  of  immune  bodies  by  unorganized  suspensions. 
W.  BiLTZ,  H.  Much  and  C.  Siebert*  have  already  called  attention 
to  the  fact  that  the  adsorption  of  their  antigens  by  hydrogels  was 
only  slightly  reversible.  Only  to  this  extent  was  J.  Bordet's*  com- 
parison of  immune  reactions  to  the  dyeing  of  fiber  with  dyes  appro- 
priate. This  irreversibility  has  its  analogies  in  the  adsorption  of 
numerous  other  known  substances  in  which  we  assume  that  secondary 
changes  occur  as  a  result  of  the  concentration  at  the  surface;  some  of 
these  changes  are  chemical,  e.g.,  the  adsorption  of  crystal  violet  and 
of  rennin  by  bone  black. 

Of  great  interest  are  the  observations  of  L.  Jacque'  and  E.  Zunz* 
illustrating  the  competing  action  of  several  adsorbents  for  a  single 
substance.  They  found  that  the  adsorption  of  diphtheria  toxin  by 
bone  black  was  reversible  in  the  body  but  irreversible  in  vitro 
[probably  because  of  protective  substances.  Tr.].  The  adsorption 
of  diphtheria  antitoxin  is,  on  the  contrary,  irreversible  in  the  body 
and  reversible  in  vitro.  Serum  albumin  may  prevent  the  adsorption 
of  diphtheria  toxin  and  antitoxin  by  bone  black. 

Adsorption  by  Organized  Suspensions. 

If  agglutinin  is  added  to  bacteria,  or  hemolysin  to  blood  corpuscles 
with  the  same  quantity  of  the  suspension,  proportionately  more  ag- 
glutinin or  hemolysin  will  be  combined  from  a  dilute  solution  than 


200 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


from  a  solution  that  is  concentrated.  Eisenberg  and  Volk*  dem- 
onstrated this  for  typhoid  bacilU  and  cholera  vibrios  and  Sv. 
Arrhenius*  and  his  co-workers  for  hemolysins  (see  also  G.  Dreyer, 
J.  Sholto  and  C.  Douglas*). 

This  is  illustrated  by  a  table  (after  Eisenberg  and  Volk)  showing 
the  combination  of  agglutinin  with  a  uniform  quantity  of  typhoid 
bacilli  and  increasingly  concentrated  agglutinin  solutions. 


Agglutinin  fixed  by  the 

Agglutinin  free  in  the 

bacteria. 

solution. 

2 

0 

20 

0 

40 

0 

i                   180 

20 

340 

60 

1,500 

500 

6,500 

3500 

11,000 

9000 

The  course  is  entirely  that  of  an  adsorption  curve. 

Cases  also  occur,  according  to  the  investigations  of  G.  Dreyer,  J. 
Sholto  and  C.  Douglas*  in  which,  after  exceeding  a  certain  maxi- 
mum, less  and  less  agglutinin  is  taken  up  by  the  bacteria,  in  spite  of 
greater  concentration  of  the  agglutinin,  typical  "abnormal  adsorp- 
tion." 

It  may  be  mentioned,  moreover,  concentrated  salt  solutions  inter- 
fere with  the  fixation  of  agglutinin  (until  now  this  had  been 
demonstrated  only  for  blood  corpuscles).  (K.  Landsteiner  and 
St.  Weleck.)  Analogous  to  this  is  an  observation  of  W.  Biltz,*^ 
according  to  which  the  addition  of  salt  interferes  with  the  adsorp- 
tion of  proteins  by  inorganic  colloids.^ 

As  has  been  stated  elsewhere,  hemolysis  with  immune  sera  occurs 
through  the  interaction  of  two  components;  amboceptor  is  bound  by 
the  blood  corpuscles  and  it  causes  the  fixation  of  the  complement 
which  accomplishes  the  hemolysis.  This  is  obviously  very  similar 
to  what  occurs  with  mordant  and  dye;  the  dye  is  fixed  to  the  fiber 
by  means  of  the  mordant.  K.  Landsteiner  and  N.  Jagic*  have  to  a 
certain  extent  devised  a  model  for  the  process;  as  amboceptor  they 
use  silicic  acid  hydrosol,  as  complement  active  serum  or  lecithin. 

Sihcic  acid  precipitates  with  blood  corpuscles  as  well  as  with  leci- 
thin; it  thus  links  together  blood  corpuscles  and  lecithin,  concentrating 

1  In  my  opinion  it  is  chiefly  globulin  which  has  been  strongly  adsorbed  in 
these  experiments,  since  the  final  portion  of  globuUn  is  separated  very  slowly  from 
dialyzed  serum. 


IMMUNITY  REACTIONS  201 

./ 
the  lecithin  on  the  surface  of  the  blood  corpuscles.  It  is  very  prolj- 
able,  as  assumed  by  the  investigators  mentioned,  that  lecithin  in  this 
instance  acts  as  a  solvent  for  the  lipoid  membrane  of  the  blood  cor- 
puscles. Numerous  similar  models  in  which  complement  was  re- 
placed by  lipoid  were  subsequently  devised. 

Reversibility.  The  combination  of  agglutinin  with  bacteria  and  red 
blood  corpuscles  is  partially  reversible;  it  may  be  partially  removed 
at  a  higher  temperature  by  washing  with  physiological  salt  solution 
(K.  Landsteiner,  Eisenberg  and  Volk). 

This  is  deduced  from  an  experiment  of  J.  Jogs.*  He  mixed  typhoid 
bacilli  bearing  agglutinin  with  untreated  typhoid  bacilli,  and  all 
the  bacilli  became  agglutinated.  There  must  have  been  a  with- 
drawal of  agglutinin  from  the  typhoid  bacilli  which  had  been  treated. 
In  a  similar  way,  J.  Morgenroth*  ^  demonstrated  that  the  combi- 
nation of  amboceptor  and  red  blood  corpuscle  is  partially  reversible. 
Reversibility  within  the  organism,  where  numerous  varieties  of  cells 
occur,  is  of  great  practical  importance. 

B.  The  Distribution  of  Immune  Substances  Between  Dis- 
solved Colloids  and  Solvent. 

The  colloid-chemical  theories  regarding  the  combination  of  toxin 
and  antitoxin  are  tacitly  based  upon  the  assumption  that  toxin  and 
antitoxin  behave  like  a  suspension  or  a  hydrogel;  they  premise  that 
surfaces  occur,  upon  which,  for  instance,  the  toxin  may  concentrate 
in  accordance  with  the  laws  of  adsorption.  Theoretical  basis  for  this 
assumption  is  lacking.  Very  little  is  known  concerning  the  fixation 
of  crystalloids  and  of  hydrosols  by  hydrosols,  when  no  precipitate 
occurs.  There  are,  at  present,  two  methods  of  attacking  the  problem. 
By  means  of  ultrafiltration,  H.  Bechhold**  has  shown  that  the  com- 
bination of  methylene-blue  with  serum  albumin  satisfies  the  conditions 
of  an  adsorption  (see  p.  25).  L.  Michaelis  and  P.  Rona  have  used 
the  osmotic  compensation  method  in  order  to  determine  the  kind  of 
combination  in  which  sugar,  Ca,  etc.,  are  fixed  in  the  blood.  Both 
methods  are  recent  and  had  not  previously  been  utilized  in  the  solu- 
tion of  this  problem.»  On  this  account  I  consider  it  unprofitable  to  dis- 
cuss at  present  the  manner  in  which  toxin  and  antitoxin  are  combined. 

It  may  be  mentioned  that  the  toxin-antitoxin  combination  is  in- 
completely reversible  in  part  and  in  part  irreversible.  P.  Ehrlich 
and  his  pupils  demonstrated  by  numerous  biological  investigations, 
that  the  combination  between  diphtheria  toxin  and  its  antitoxin 
rapidly  became  irreversible. 

The  relations  between  precipitin  and  precipitable  substance  is  some- 


202  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

what  more  obvious.  Inasmuch  as  the  two  dissolved  colloids  yield 
a  precipitate  when  mixed  in  suitable  proportion,  we  can  form  a 
judgment  concerning  the  proportionate  quantities  that  combine. 
However,  this  has  regard  for  the  composition  after  precipitation.  Ac- 
cording to  E.  VON  Düngern*  the  precipitate  binds  much  more  precip- 
itin than  is  required  to  cause  complete  precipitation.  It  is  still  a 
question  whether  this  combination  existed  in  the  solution. 

C.  Precipitation  of  Dissolved  Colloids  and  Organized 
Suspensions, 

Serum  containing  precipitin,  for  instance,  goat-rabbit  serum, 
gives  a  precipitate  with  its  antigen  (goat  serum).  The  serum  is 
precipitated  by  the  precipitin  just  as  it  would  be  hy  an  inorganic 
hydrosol  or  an  acid  protein  (histone),  U.  Friedemann  and  H.  Frieden- 
thal* (see  p.  157).  The  precipitation  occurs  best  in  the  presence  of 
an  optimum  mass  proportion  between  precipitin  and  precipitable 
substance;  excess  of  precipitable  substance  interferes  with  the  pre- 
cipitation. A  precipitation,  according  to  M.  Neisser,*  occurs  also 
in  salt-free  solution  (which  contains  no  globulin)  but  the  precipi- 
tation zone  differs  from  that  in  a  solution  containing  salt. 

Though  the  mutual  precipitation  of  two  amphoteric  colloids  de- 
pends on  the  hydrogen  ion  concentration  (see  p.  147),  specific  precip- 
itations (this  applies  to  precipitins  and  agglutinins)  are  largely  in- 
dependent of  it  (L.  Michaelis  and  Davidsohn).  The  electric  charge 
of  the  components  plays  a  very  subordinate  part  in  these  precipitations. 

The  plant  toxins,  ricin  (from  the  seeds  of  varieties  of  castor  bean) 
and  abrin  (from  jequirity  seeds)  have  a  similar  precipitating  effect 
upon  albumin. 

The  conditions  in  the  case  of  organized  bacterial  albumin  are 
similar  to  (but  not  identical  with)  those  of  serum  albumin.  If  an 
animal  (e.g.,  a  rabbit)  is  injected  with  bacteria  (for  instance,  typhoid 
bacilli)  there  develops  in  its  blood  an  agglutinin  which  causes  typhoid 
bacilh  in  a  test  tube  to  precipitate.^  Agglutinin  forms  a  compound, 
with  the  (actual)  albuminous  capsule  of  the  bacteria,  so  that  these 
behave  as  though  they  were  changed  from  a  hydrophile  to  a 
hydrophobe  suspension.  Precipitation  occurs  only  in  water  con- 
taining salt.2    Though  a  suspension    of   bacteria  is  unchanged  by 

^  This  phenomenon  was  first  observed  by  Gruber  and  Durham.  Widal  was 
the  first  to  use  it  for  diagnosis,  and  since,  as  the  Gruber-Widal  Reaction,  it  is  em- 
ployed in  the  diagnosis  of  typhoid,  paratyphoid,  dysentery,  etc. 

^  According  to  U.  Friedemann  it  is  possible  to  obtain  agglutination  in  a  salt- 
free  solution,  though  this  has  nothing  in  common  with  specific  agglutination. 
The  resemblance  to  the  precipitins  is,  in  this  respect,  only  a  superficial  one. 


IMMUNITY  REACTIONS 


203 


dilute  alkali  salts,  these  ^alts  cause  a  precipitation  of  agglutinated 
bacteria  as  they  would  a  suspension  of  kaolin  or  mastic.  This  was 
demonstrated  by  H.  Bechhold*^  as  well  as  M.  Neisser  and  U. 
Fribdemann*,  and  practically  confirmed  by  B.  H.  Buxton,  P. 
Shaffer  and  0.  Teague*,  as  well  as  by  B.  H.  Buxton  and  A.  H. 
Rahe*. 

Capsulated  bacteria  possess,  in  their  mucous  capsules,  a  natural 
protective  colloid  and  accordingly,  they  are  not  agglutinated  by 
immune  serum  even  in  suspensions  containing  salt.  0.  Forges 
showed  that  if  the  mucous  capsules  were  removed  by  gently  heating 
them  with  dilute  hydrochloric  acid,  even  capsulated  bacteria  were 
agglutinated  by  immune  sera. 

As  in  the  case  of  the  precipitins,  a  certain  quantitative  rela- 
tionship between  bacteria  and  agglutinating  serum  is  required  for 
precipitation.  But  in  this  instance,  there  are  certain  irregularities 
as  regards  native  and  heated  sera. 

This  phenomenon  also  has  its  analogue  in  the  precipitation  of 
unorganized  suspensions  in  the  presence  of  protective  colloid.  Table  I 
(see  below)  illustrates  the  agglutination  of  bacteria  by  diluted  immune 
serum.  Table  II  illustrates  the  precipitation  of  a  mastic  suspension 
by  AI2 (804)3  in  the  presence  of  leech  extract  as  a  protective  colloid. 

These  "irregular  series"  (see  p.  84)  frequently  occur  in  the  pre- 
cipitation of  suspensions  by  ferric  chlorid,  aluminium  chlorid  and 
certain  dyes.  They  are  explained  by  the  fact  that  the  hydrolytically 
split  iron  oxid  hydrosol,  etc.,  functions  as  a  ''protective  colloid,"  and 
in  certain  proportions  interferes  with  the  precipitation.  If  still  an- 
other protective  colloid  (gelatin,  leech  extract,  etc.)  is  added,  the 
circumstances  are  still  further  complicated  as  shown  by  Table  II. 


TABLE  I. 


TABLE  II. 


Agglutination  of  typhoid  bacilli  with 
very  dilute  immune  serum.     (After 
Eisenberg  and  Volk.) 

Precipitation    of    mastic    suspension 
by   0.0002    c.c.  J  Al2(S04)3   m  the 

presence  of   leech  extract    as  pro- 
tective  colloid.      (After  M.  Neis- 
ser and  U.  Friedemann,  and  H. 
Bechhold.) 

Dilution  of 
the  serum. 

Appearance  after  2  hours. 

Dilution  of 
leech  extract. 

Appearance  after  24  hours. 

1/100 
1/1000 

1/25000 
1/30000 
1/45000 

No  a.gglutination 
Almost  complete  agglu- 
tination 
Trace 

Heavy  flocks 
No  agglutination 

c.c. 
0.1 

0.03 

0.01 

0.003 

0.001 

No  precipitation 
Almost    complete    pre- 
cipitation 
No  precipitation 
No  precipitation 
Complete  precipitation 

204  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Since  bacteria,  like  other  proteins,  may  be  precipitated  by  acids 
without  previous  treatment  with  agglutinating  serum,  L.  Michae- 
lis and  Beniasch  tested  different  groups  of  bacteria  (typhoid,  para- 
typhoid, colon,  etc.)  to  determine  whether  the  result  depended  on 
the  H  ion  concentration.  They  found  that  different  concentrations 
of  H  ions  were  necessary  for  the  precipitation  of  different  groups  of 
bacteria.  The  discoverers  based  on  this  fact,  a  method  for  distin- 
guishing certain  bacterial  groups.  It  is  still  a  question  whether  the 
procedure  is  practical  either  alone  or  in  combination  with  agglutinat- 
ing serum  (see  S.  Galitzer). 

Blood  corpuscles,  resembling  bacteria,  behave  like  a  hydrophile 
suspension  whose  surface  is  so  changed  by  various  agglutinins  that 
they  flock  out.  I  am  inclined  to  believe  that  a  true  glueing  together 
occurs  more  frequently  with  blood  corpuscles  than  with  bacteria. 

From  our  previous  experiments,  we  see  that  colloids  and  suspen- 
sions are  precipitated  not  only  by  electrolytes  but  also  hy  colloids  of 
opposite  charge.  It  must  therefore  be  possible  to  agglutinate  or- 
ganized suspensions  as  bacteria  and  blood  corpuscles  by  suitable 
hydrosols.  Experiments  with  hydrosols  of  iron,  zirconium,  thorium 
oxid  and  silicic  acid  ^  (W.  Biltz,  H.  Much  and  C.  Siebert*,  also  K. 
Landsteiner  and  N.  Jagic*,  Girard-Mangin  and  V.  Henri*)  con- 
firm this  assumption  and  show  that  as  with  other  colloid  precipi- 
tations in  salt  solutions,  an  optimum  proportion  between  the  two 
colloids  must  exist,  or  no  precipitation  will  occur. 

It  should  be  emphasized  that  the  combination  of  bacteria  or  blood 
corpuscles  and  inorganic  hydrosols  is  irreversible  (in  contradistinction 
to  the  agglutinin  combination). 

Blood  corpuscles  differ  very  greatly  in  one  respect  from  bacteria. 
Though  the  latter  migrate  to  the  anode  showing  their  negative 
charge,  blood  corpuscles  are  more  amphoteric.  As  a  result  of  this, 
they  are  precipitated  by  negative  hydrosols  (arsenic  tri-sulphid, 
silicic  acid,  etc.)  L.  Hirshfeld*^.  A  very  important  observation  is 
that  of  L.  Hirshfeld,  that  the  agglutination  of  blood  corpuscles  of 
different  animals  by  zinc  nitrate  follows  the  same  order  of  precipita- 
bility  as  their  agglutination  by  agglutinating  sera  and  abrin. 

Ricin  and  abrin  also  agglutinate  blood  corpuscles,  obviously,  be- 
cause they  precipitate  their  albumin. 

From  our  entire  exposition,  it  is  evident,  that  the  adsorption  of  the 
agglutinating  substance  and  the  agglutination  are  two  separate  proc- 
esses which,  in  their  principle,  have  nothing  in  common.  The  agglu- 
tinin changes  the  bacteria  or  erythrocytes,  making  them  agglutinable; 

1  Colloidal  silicic  acid  agglutinates  in  much  greater  dilution  than  the  crys- 
taUoidal. 


IMMUNITY  REACTIONS  205 

the  electrolyte,  which  itself  is  not  adsorbed,  agglutinates  or  flocks 
them  out.i  It  is  therefore  obvious  that  an  electrolyte  which  changes 
the  cells  may,  nevertheless,  without  being  adsorbed,  agglutinate  them. 
According  to  J.  Dunin-Borkowski*,  red  blood  corpuscles  are  agglu- 
tinated by  FeCls,  though  it  is  not  combined  with  them. 

Electric  Charge,  H  and  OH  Ions. 

Numerous  attempts  have  been  made  to  determine  the  electric 
charge  of  antigens  and  immune  substances  by  cataphoresis  (K. 
Landsteiner  and  Wo.  Pauli*,  C.  N.  Field  and  0.  Teague*.  It  is 
so  small,  however,  that  in  my  opinion  it  cannot  be  definitely  rec- 
ognized since  traces  of  H  or  OH  ions  may  cause  a  reversal  of  charge 
(see  Bechhold*^*'). 

This  also  applies  to  the  results  of  exhaustive  adsorption  experiments 
with  adsorbents  of  opposite  electric  charge,  namely,  the  experiments 
carried  out  by  Edgar  Zunz  with  electro-osmotically  purified  sihcic 
acid,  aluminium  hydroxid,  kaolin,  diatomaceous  earth,  talc,  and  clay 
upon  toxins  and  antitoxins.  With  K.  Landsteiner  and  W.  Pauli  it 
is  well  to  regard  antigens  and  antibodies  as  amphoteric  electrolytes. 

The  difference  between  bacteria  and  bacteria  bearing  agglutinin 
is  clearly  demonstrated  (H.  Bechhold*i,  M.  Neisser  and  U.  Friede- 
mann*). Though  the  former  (typhoid,  dysentery)  migrate  to  the 
anode,  the  latter  lose  their  charge  on  account  of  the  agglutinin  and 
precipitate  between  the  electrodes. 

More  characteristic  than  electrical  migration  is  the  behavior  of 
toxins,  antigens  and  immune  bodies  to  acids  and  alkahs.^  From 
our  standpoint,  only  very  weak  dilutions  of  H  and  OH  are  considered 
from  such  as  cause  no  irreversible  destruction  in  the  substances 
affected.  Acidity  diminishes  the  toxicity  of  some  toxins,  but  it  is 
restored  by  neutraHzing  them.  Kirschbaum  isolated  a  nontoxic 
dysentery  toxin  by  ultrafiltration  and  precipitation  with  acids;  it 
dissolved  in  alkalis  and  forms  a  toxic  salt-like  combination.  An 
observation  made  by  J.  Morgenroth  *2  points  to  the  occurrence  of  a 
salt-like  combination  of  cobra  hemolysin  and  also  of  cobra  neuro- 
toxin; these  neutral  toxins,  although  colloids,  diffuse  through  an 
animal  membrane  into  a  solution  containing  hydrochloric  acid. 
Cobra  venom  may  be  recovered  from  crystalloid  cobra  venom  salt 
by  neutralizing  it,  yet  it  gradually  goes  into  the  colloidal  state  as  I 

1  P.  Schmidt  assumes  an  additional  phase;  modification  of  the  bacteria  by 
agglutinin,  adsorption  of  globulin  by  modified  bacteria,  flocculation  by  elec- 
trolytes. 

2  Only  great  dilutions  of  H  and  OH  are  here  considered,  such  as  do  not 
cause  an  irreversible  change  in  the  material. 


206  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

have  inferred  from  the  following  experiments  of  J.  Morgenroth  and 
D.  Pane.*5 

I  wish  to  call  attention  to  one  other  property  which  is  strongly 
suggestive  of  colloids.  J.  Morgbnroth  and  D.  Pane*  heated 
cobra  venom  in  n/20  HCl  solution  and  determined  its  hemolytic 
action  immediately  after  cooling  and  neutraUzation.  The  hemolytic 
activity  induced  by  lecithin  was  greatly  diminished  but  gradually 
(after  hours  or  days)  resumed  its  original  strength.  It  seems  reason- 
able to  regard  the  gradual  restoration  of  toxicity  as  phenomena  of 
"maturation"  since  the  particles  of  the  molecular  dispersed  cobra 
hemolysin,  gradually  unite  to  larger  agglomerations  and  thus  ac- 
quire greater  adsorptive  capacity. 

Colloid  chemistry  offers  numerous  similar  examples;  to  mention 
only  the  aging  of  dye  solutions  (hemotoxylin)  which  must  occur  pre- 
vious to  its  utilization  in  histological  stains.  The  same  interpreta- 
tion applies  to  the  anologous  observation  upon  the  neurotoxin  of 
cobra  venom. 

As  a  rule  the  union  of  antigen  and  immune  substances  is  inhibited 
both  by  H  and  by  OH  ions.  Just  as  H  and  OH  ions  may  break  down 
the  union  of  toxin  and  antitoxin  so  may  they  dissolve  the  bonds 
holding  agglutinin  to  its  substrate  (Hahn  and  R.  Trommsdorf),  or 
either  abrin  or  amboceptor  to  blood  corpuscles. 

The  influence  of  reaction  on  the  action  of  hemolytic  sera  is  told  in 
the  researches  of  S.  Abramow,  Hecker,  L.  von  Liebermann,  P. 
RoNDONi,  H.  Sachs  and  Altmann,  L.  Michaelis  and  Skwirsky  (see 
P.  RoNDONi*  for  bibliography). 

Under  certain  conditions  hemolysis  is  hastened  by  slight  acidity 
and  retarded  by  larger  quantities  of  acids  or  by  alkahs. 

The  inhibiting  action  of  alkah  and  to  a  less  definite  degree,  of  acid, 
is  evident  in  antigen-antibody  combinations,  as  is  revealed  by  com- 
plement deviation  (see  p.  207).  Its  significance  is  also  evident  in  the 
Wassermann  reaction. 

Addition  of  1/1000  to  1/3200  normal  NaOH  may  inhibit  the  reac- 
tion in  a  strongly  positive  serum;  similarly,  a  negatively  reacting 
(luetic  serum)  may  give  a  strongly  positive  reaction  after  the  addition 
of  1/1000  to  1/2000  HCl  (H.  Sachs  and  Altmann). 

The  following  findings  favor  the  view  that  the  physical  fixation  of 
amboceptor  by  blood  corpuscles  is  influenced  by  the  reaction.  By 
means  of  alkali,  the  blood  corpuscles  may  be  prevented  from  com- 
bining with  the  amboceptor;  on  the  other  hand  amboceptor-laden 
blood  corpuscles  may  be  deprived  of  amboceptor  by  alkahs,  and 
the  amboceptor  may  be  recovered  in  an  active  condition.  The  facts 
for  acids  are  not  so  obvious. 


IMMUNITY  REACTIONS  207 


Complement  Fixation  and  the  Wassermann  Reaction. 

A  mixture  of  antigen  and  immune  substance  {e.g.,  goat  serum  + 
goat-rabbit  serum)  has  the  property  of  binding  complement.  This  is 
recognized  by  the  following:  if  complement  is  added  to  a  suspension 
of  red  blood  corpuscles  +  amboceptor,  hemolysis  occurs.  If  the 
complement  has  previously  been  mixed  with  antigen  +  immune 
substance  and  we  then  add  the  entire  mixture  to  the  blood  corpuscles 
+  amboceptor,  no  hemolysis  occurs. 

Complement  Complement 

+  + 

Amboceptor  Amboceptor       Amboceptor       +         Antigen 

+  +  +  + 

Erythrocytes  Erythrocytes      Erythrocytes  Immune  substance 


Hemolysis  No  Hemolysis  No  Hemolysis 

This  phenomenon,  which  is  called  com'plement  fixation  or  com- 
plement deviation,  was  discovered  by  0.  Gengou  and  Bordet.  It 
has  acquired  great  practical  significance  by  its  utilization  for  the 
recognition  of  antigen  by  M.  Neisser  and  H.  Sachs  (one  billionth 
c.c.  of  human  blood  may  be  recognized  by  complement  deviation) 
and  also  indirectly,  through  a  reaction  analogous  to  the  Wassermann 
reaction  for  the  recognition  of  luetic  infection. 

A  mixture  of  antigen  and  immune  serum  may  give  a  precipitate, 
though  only  when  mixed  in  definite  proportions,  otherwise  this  does 
not  occur.  Complement  fixation  occurs  regardless  of  the  occurrence 
of  a  precipitation.  Since  complement  is  easily  adsorbed  by  many 
colloids  and  suspensions,  it  was  natural  to  suppose  that  the  precipi- 
tate of  antigen  and  immune  substances  was  the  fixing  agent.  This 
view  is  held  especially  by  U.  Friedemann  who  attributes  to  eu- 
globulin  the  complement  binding  power  of  the  immune  serum.  The 
investigations  of  Dean  are  of  great  interest  to  students  of  colloid 
chemistry.  According  to  these  investigations,  much  complement 
is  bound  when  there  is  a  slowly  developing  turbidity  and  but  Kttle 
when  turbidity  develops  rapidly.  From  this  aspect  a  definite  develop- 
ment of  surface  tension  favors  binding  of  the  complement.  Though 
it  is  probable  that  binding  of  complement  depends  on  a  physical  ad- 
sorption of  the  visible  or  invisible  precipitate,  its  mechanism  requires 
further  elucidation.  The  objection  that  the  fixation  may  occur  even 
without  the  appearance  of  a  precipitate  cannot  definitely  be  proven. 
We  know  that  albumin  particles  may  aggregate  into  larger  particles 
without  a  precipitation,  provided  the  excess  of  one  of  the  precipitate- 
forming  colloids  acts  as  a  protective  colloid.     On  the  other  hand,  it 


208  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

has  not  yet  really  been  demonstrated  that  a  physical  fixation  and 
not  an  irreversible  chemical  change  occurs  in  complement  fixation. 
Therefore,  in  what  category  complement  fixation  by  means  of  anti- 
gen plus  immune  substance  is  to  be  placed,  is  still  an  open  question. 


Wassermann  Reactjon. 

Complement  fixation  is  much  more  obvious  in  the  Wassermann 
reaction.  When  A.  Wassermann  began  his  studies,  he  started  with 
the  assumption  that  extract  of  spirochsete  (as  antigen)  -|-  luetic 
serum  (as  immune  substance)  must  fix  complement.  It  was  very  soon 
evident,  that  spirochsete  extract  could  be  replaced  by  numerous 
lipoid  suspensions: — by  lecithin  (0.  Purges  and  Maier),  by  sodium 
glycocholate  (Levaditi  and  Yamanouchi),  vaseline  (Fleischmann), 
by  sodium  oleate  (H.  Sachs  and  Altmann),  sodium  palmitate  and 
stearate  (P.  Hessberg),  potato  extract  and  emulsion  of  shellac 
(Munk).  As  Elias,  0.  Neubauer,  0.  Purges  and  Salomon 
showed,  these  hydrophile  lipoids  give  precipitates  with  the  globulin 
of  luetic  sera  which  fix  complement.^ 

The  parallelism  between  precipitation  reaction  and  complement 
fixation  is  very  suggestive  of  a  colloid  phenomenon.  It  would  be 
a  convincing  proof  that  complement  fixation  in  the  Wassermann 
reaction  was  a  surface  phenomenon,  if  it  could  be  demonstrated 
that  the  reaction  did  not  occur  in  the  absence  of  a  precipitation. 
This  proof  I  gather  from  an  observation  of  H.  Sachs  and  P.  Rondoni. 
They  found  that  complement  fixation  by  an  alcoholic  extract  of  syph- 
ihtic  livers^  depended  upon  the  manner  of  dilution  with  saline  solu- 
tion; diluting  drop  by  drop,  they  obtained  a  fluid  which  bound 
complement  strongly.  If  the  extract  is  added  rapidly  to  the  saline 
solution,  there  occurs  a  weak  complement  fixation  or  none  at  all. 
By  slowly  adding  drop  by  drop  we  obtain  a  turbid  fluid;  by  rapid 
mixing  a  clear  fluid.  We  have  here  two  fluids,  which  in  their  ability 
to  fix  complement  can  be  distinguished  only  hy  the  surfaces  of  the 
suspended  lipoids.  The  observation  of  F.  Munk  also  confirms  this 
view  that  only  alcoholic  or  acetone  and  not  ethereal  extracts  or 
solutions  of  the  above-mentioned  lipoids  are  suitable  for  binding  of 
complement. 

1  The  observations  of  these  authors  are  very  interesting.  They  observed  that 
even  normal  sera  give  precipitates  with  the  hpoids,  but  that  the  range  of  preci- 
pitation is  much  broader  with  luetic  sera  and  that  the  complement  fixation  pre- 
supposes an  optimum  mass  relationship  of  lipoid  and  serum. 

2  Before  it  was  discovered  that  the  above-mentioned  lipoids  were  suitable  for 
the  Wassermann  reaction,  extracts  of  syphilitic  livers  were  employed. 


IMMUNITY  REACTIONS  209 

P.  Hessberg  observed^  that  freshly  prepared  solution  of  sodium 
palmitate  bound  complement,  but  it  lost  this  property  by  repeated 
heating.  This  change  by  means  of  repeated  heating  is  a  character- 
istic colloid  property,  which  evidently  is  associated  with  a  fragmen- 
tation of  the  particles;  the  more  frequently  gelatin,  agar-agar,  etc., 
are  heated,  the  more  difficult  it  is  to  solidify  them.  Unfortunately 
P.  Hessbeeg  did  not  determine  whether  the  repeatedly  heated  solu- 
tion of  sodium  palmitate  recovered,  on  long  standing,  its  ability  to 
fix  complement. 

ANAPHYLAXIS,   DEFENSIVE   FERMENTS,   AND   MEIO- 
STAGMIN   REACTION. 

Anaphylaxis. 

If  an  animal  {e.g.,  a  guinea  pig)  is  injected  with  antigen  (e.g.,  horse 
serum)  there  are  no  sequelae.  If  the  injection  is  repeated  after  an 
interval  of  about  10  to  14  days  there  occur  serious  symptoms  of  poison- 
ing (convulsions,  rise  of  temperature  and  respiratory  distress)  which 
frequently  terminate  fatally  in  a  few  minutes  (anaphylactic  shock). 
This  condition  induced  by  the  first  injection  of  serum  or  bacteria  is 
called  anaphylaxis  (induced  defenselessness  —  the  reverse  of  im- 
munity). Human  "serum  sickness"  is  also  a  phenomenon  of  ana- 
phylaxis which  appears  after  the  repeated  injection  of  curative  sera 
and  manifests  itself  in  erythemata,  swelling  of  the  lymph  nodes  and 
joints  and  moderate  rises  of  temperature.  Anaphylaxis  is  strongly 
specific,  which  means  that  it  occurs  only  upon  repeated  injections  of 
the  same  protein  or  the  same  strain  of  bacteria.  The  specificity  is 
so  absolute  and  the  quantities  required  so  minute,  that  like  precipita- 
tin  reactions,  anaphylactic  phenomena  may  be  employed  in  dis- 
tinguishing traces  of  human  from  animal  blood  or  in  detecting 
adulterations. 

Friedberger  and  his  coworkers  were  successful  in  preparing  the 
anaphylactic  poison  (anaphylatoxin)  outside  the  body,  in  vitro.  If 
an  animal  (e.g.,  a  guinea  pig)  is  injected  with  antigen  (e.g.,  horse 
serum)  antibody  appears  in  the  blood  after  a  time.  Friedberger 
proceeded  from  this  fact;  he  mixed  antigen  and  antibody  in  a  test 
tube  and  obtained  a  precipitate  from  the  mixture.  By  digesting 
the  precipitate  with  guinea-pig  serum  (which  always  contains  com- 
plement) he  obtained  what  he  designated  as  anaphylotoxin. 

Since  peptones  produce  phenomena  resembhng  anaphylactic  shock, 
it  was  thought  that  peptone-like  products  were  split  off,  perhaps  by 
fermentation  in  the  interaction  between  antibodies  and  antigen. 


210  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

This  view,  still  held  by  Friedberger,  regards  the  appearance  of  the 
anaphylactic  poison  as  the  result  of  a  fermentation  of  antigen  by 
the  degradation  action  of  complement  resulting  from  antibody 
fixation. 

It  was  a  surprise  to  find  that  the  anaphylatoxin  might  be  prepared 
even  without  antibodies  as  when  guinea-pig  serum  was  digested  with 
bacteria;  and  even  by  digestion  with  colloidal  carbohydrates  (agar, 
starches,  starch  paste,  pectin  and  inulin).*  [The  work  of  Vaughan 
and  NovY  on  the  protein  poisoning  should  be  mentioned  as  well  as 
that  of  R.  Weil  on  the  mechanism  of  anaphylaxis.     Tr.] 

H.  Sachs  and  E.  Nathan  demonstrated  that  the  physical  state  of 
the  poison-producing  substances  was  a  determining  factor.  They 
employed  inulin  as  absorbent.  This  carbohydrate  is  practically 
insoluble  in  cold  water  though  it  forms  a  clear  solution  without 
pasting  in  warm  water.  Upon  mixing  guinea-pig  serum  with  a  5  per 
cent  suspension  of  inulin,  a  serum  was  obtained  which  produced  the 
severest  anaphylactic  shock  when  injected  into  guinea  pigs,  though 
no  toxic  substance  resulted  from  the  mixture  with  inulin  solution. 
Pastes  have  a  very  extensive  surface  development,  consequently, 
anaphylatoxin  is  formed  best  with  starch  paste.  H.  Sachs  and  E. 
Nathan  find  in  these  experiments  decisive  confirmation  of  the 
physical  theory  of  anaphylaxis  first  proposed  by  H.  Sachs  and  Ritz. 
According  to  them,  antigen  is  not  the  mother  substance  of  anaphyla- 
toxin, which  is  not  newly  formed,  but  which  exists  preformed  in 
normal  serum.  This  poison  becomes  active  by  the  adsorption 
(separation)  of  a  substance  as  yet  undetermined.  In  the  case  of 
artificial  anaphylatoxins,  bacteria  and  carbohydrates  serve;  in  true 
anaphylaxis,  the  products  of  antigen  and  immune  body  act  as  ad- 
sorbents« The  specificity  of  anaphylaxis  is  thus  explained  since  only 
the  specific  antibody  formed  causes  a  orecipitate  with  the  antigen. 

Protective  Ferment. 

The  remarkable  relation  between  immunity  reactions  and  pro- 
tective ferments  should  be  mentioned  here.  E.  Abderhalden  con- 
siders under  this  term  the  ferments  which  destroy  and  render 
innocuous  species-foreign  proteins,  entering  the  organism  parent- 
ally. Since  the  connection  between  protective  ferments  and  colloid 
research  is  still  unestablished  we  shall  merely  refer  to  the  work  of 
E.  Abderhalden.^ 

1  It  is  not  yet  definitely  established  that  anaphylatoxin  may  be  made  by 
shaking  inorganic  suspension  with  normal  guinea-pig  serum. 

2  [D.  Van  Slyke  has  thoroughly  discredited  the  Abderhalden  reaction  with  his 
nitrous  acid  method  for  the  quantitative  determination  of  amino  acids.  Harvey 
Lectures,  1915-16,  p.  170,  Lippincott,  N.  Y.    Tr.J 


IMMUNITY  REACTIONS  211 

The  Meiostagmin  Reaction. 

M.  AscoLi  and  G.  Izar  found  that  substances  which  lower  surface 
tension  of  the  solution  are  formed  in  the  reaction  between  antigens 
and  immune  bodies.  He  determined  this  with  Traube 's  stalag- 
mometer  by  counting  the  drops  which  formed  from  a  measured 
quantity  of  fluid.^  When,  for  instance,  he  mixed  an  extract  of 
typhus  bacilli  with  normal  serum  and  with  the  serum  of  typhoid 
patients,  10  c.c.  gave  58  drops  in  the  former  instance  and  61  drops  in 
the  latter.  M.  Ascoli  considers  it  to  be  a  general  reaction  and  has 
employed  it  in  the  serum  diagnosis  of  various  conditions  (syphiHs, 
tuberculosis,  anchylostomiasis  and  echinococcus  infection).  It  has 
not  been  generally  introduced  as  a  means  chnical  diagnosis  of  infec- 
tious disease  since  the  technic  is  so  precise  that  the  differences  are 
within  the  limit  of  error,  but  it  has  been  more  frequently  employed 
in  the  detection  of  malignant  growths.  Two  dilutions  of  the  serum 
are  prepared,  one  with  water  and  the  other  with  an  equal  quantity 
of  tumor  extract  (recently  Ascoli  employed  ricinoleic  acid  or  linoleic 
acid,  etc.).  If  the  number  of  drops  are  larger  in  the  latter  mixture 
than  in  the  former,  there  is  a  presumption  that  the  serum  is  from 
a  cancer  patient.^ 

1  Meiostagmin.  reaction  =  reaction  of  smaller  sized  drops. 

2  [E.  P.  Bernstein  and  Irving  E.  Simons,  after  critically  reviewing  the  litera- 
ture and  their  personal  experience,  have  discarded  the  Meiostagmin  reaction  as 
useless  cUnically.    Amer.  Jour.  Med.  Sei.,  vol.  142,  p.  862,  et  seq.     Tr.] 


An  asterisk  (*)  after  an  author's  name  refers  to  a  reference  in  the  index  of 
names. 

PART   III. 

THE    ORGANISM   AS   A   COLLOID    SYSTEM. 

The  Significance  of  the  Colloidal  Condition  for  the  Organism. 

Recently,  I  read  in  a  French  magazine,  a  fantastic  description 
of  a  visit  to  the  Martians.  They  were  pictured  as  men  mth  iron 
faces;  with  a  great  bill  replacing  the  nose;  three  glass  eyes  and 
joints  and  limbs  of  iron.  VThj  did  not  the  artist  construct  his 
people  of  a  material  actually  found  on  that  planet?  If  we  assume 
that  life  exists  on  other  planets  and  disregard  the  theory  of  pan- 
spermogenesis  ^  accepted  as  probable  by  Sv.  Aerhexius,  a  priori  it  is 
probable  that  life  is  associated  with  substances  similar  to  those  with 
which  it  is  associated  on  our  earth.  One  thing  I  can  assert,  that 
whatever  the  material  comyosition  of  such  liimig  beings  may  he,  it  must 
he  colloidal  in  nature. 

Those  iron  Martians  could  no  more  exist  than  could  crystallized 
life.  What  condition  of  matter,  other  than  the  colloidal,  could 
adopt  such  changeable,  such  plastic  shapes,  and  yet,  when  necessary, 
be  in  a  position  to  maintain  them. 

An  exchange  of  substance  may  occur  in  jellies  as  well  as  in  a  fluid; 
in  the  latter  the  least  touch,  an  miintentioned  movement,  disturbs 
the  result  of  diffusion  and  brings  about  the  death  of  the  system; 
the  changes  in  a  jell 3"  are  fixed  as  m  a  solid  mass.  Colloids  may  form 
permeable  walls  or  membranes,  whose  permeability  is  regulated  by 
the  substances  which  pass  through  them;  thus,  for  instance,  the 
sulphates  which  are  less  important  to  the  organism  close  the  passages 
on  themselves;  the  chlorids  facilitate  their  otsti  entrance. 

Foods  enter  our  digestive  tract  in  colloidal  condition,  as  albumin 
and  starches.  Made  fluid  and  easily  diffusible  bj^  the  enzjTnes,  they 
penetrate  the  organism  in  order  to  be  fixed  and  again  transformed 
into  colloids.  In  that  condition  only  are  they  retained  by  the  or- 
ganism and  prevented  from  flowdng  away.     Colloids,  because  of  their 

^  According  to  this  theory,  it  is  conceivable  that  germs  of  life  travel  from  one 
planet  to  another  and  that  they  develop  there  under  favorable  circumstances, 
so  that,  to  a  certain  extent,  one  star  infects  another  one  with  life. 

213 


214  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

surface  development,  unite  the  advantages  of  the  soHd  condition  with 
that  of  the  fluid;  observe  for  a  moment  a  mountain  cUmber,  a  fever 
patient,  or  a  tree  in  the  springtime,  which  is  decked  with  leaves 
in  four  or  five  days.  What  enormous  amounts  of  chemical  energy 
are  expended  by  the  mountain  climber  in  a  few  hours,  what  large 
quantities  of  protein  are  in  a  short  time  consumed  by  the  fever  patient, 
what  large  quantities  of  material  are  carried  to  and  from  the  periph- 
ery of  the  tree.  With  the  least  loss  of  time  the  reserves  must  be 
mobilized  and  carried  to  the  seat  of  war,  the  places  where  they  are 
consumed.  Such  a  rapid  mobilization  is  unthinkable  in  the  case  of 
a  solid  crystalloid,  with  its  small  surface;  a  chemical  process  in  a 
swollen  colloid  may  only  occupy  minutes,  whereas  the  same  reaction 
in  a  shrunken  colloid  requires  days. 

How  wonderful  by  means  of  adsorption  is  the  action  of  surface 
development  as  a  regulating  mechanism.  Luxus  consumption  of  foods, 
salts,  oxygen,  etc.,  are  excreted  as  quickly  as  possible  by  the  colloid 
components  of  the  body,  but  when  the  supply  ceases,  the  amount 
given  off  becomes  less  and  when  there  is  a  deficiency  the  organism 
tenaciously  retains  the  last  traces  for  its  time  of  need. 

Quantitatively,  the  substance  most  important  for  the  organism  is 
water;  colloid  and  water  are  one  in  the  organism;  an  organism 
without  water  is  lifeless.  We  can  imagine  such  an  intimate  and 
varying  relation  to  water  only  in  a  colloid  system;  the  process  of 
swelling,  the  adsorption  of  water,  and  shrinking  to  complete  dryness 
exhibit  no  leaps  or  sudden  changes  in  condition.  If  we  compare 
crystalloids  with  colloids,  we  shall  see  that  something  entirely  new 
with  very  changed  properties,  a  solid  crystalloid  precipitate,  appears 
from  a  solution  upon  losing  water.  Such  a  system  would  be  unable 
to  maintain  correctly  the  constantly  oscillating  water  balance  and 
the  normal  condition  of  swelling  in  the  organism,  or  to  act  as  an 
accumulator  of  large  quantities  of  water,  like  muscle,  and  release  it 
for  use  when  necessary.  Such  a  system  could  not,  like  a  pen,  smooth 
out  the  irregular  chemical  impulses,  which  the  organism  experiences 
as  the  result  of  physiological  and  pathological  life  processes,  and 
which,  after  absorption,  constantly  restores  its  state  of  swelling  to 
normal  by  means  of  secretion  (kidney,  skin,  etc.). 

We  thus  see  that  the  processes  which  cause  us  to  marvel  at  the 
wonderful  adaptability  of  Nature  rest  upon  the  simple  laws  applicable 
to  colloids.  Thus  it  is  that  I  am  unable  to  imagine  that  the  com- 
plicated and  adaptive  phenomena  of  Life  could  possibly  be  associated 
with  any  other  than  a  colloidal  system. 


CHAPTER  XIV. 

METABOLISM   AND   THE   DISTRIBUTION    OF   MATERIAL. 

The  Distribution  of  Water  in  the  Normal  Organism. 

The  earliest  stages  in  the  development  of  life  are  accompanied  by 
powerful  processes  of  swelling  which  soon  reach  a  maximmn,  and 
then  pass  over  into  a  shrinking,  which  becomes  progressively  greater, 
until  death  occurs.  In  the  cases  of  plants,  the  struggle  for  water  be- 
tween seed  and  soil  starts  with  germination  (A.  Muntz*).  Growing 
and  full-grown  plants  show  a  certain  turgor,  i.e.,  a  fulness  or  tension 
like  a  distended  rubber  balloon,  while  a  dying  plant  is  withered  and 
poor  in  water. 

A  three  months'  human  fetus  contains  94  per  cent  water;  at  birth 
the  water  content  is  from  69  to  66  per  cent;  in  adult  life  58  per  cent.^ 
I  am  not  acquainted  with  any  figures  of  the  water  content  of  the  aged, 
but  it  is  generally  and  with  justice  assumed,  that  in  old  age,  the 
water  content  decreases;  turgescence  in  general,  and  of  the  skin  in 
particular,  is  obviously  lost.  The  organism,  taken  as  a  whole,  dur- 
ing its  life  evidently  passes  through  the  curves  of  swelling  and  shrink- 
ing of  an  inelastic  gel.  In  individuals  of  the  same  species,  the  water 
content  is  probably  fairly  constant  for  the  same  period  of  life. 

The  water  content  of  the  individual  portions  of  plants  and  of  simi- 
lar organs  of  different  plants  varies  remarkably.  Though  jelly-like 
protoplasm  contains  from  60  to  90  per  cent  of  water,  the  dry  wall  of 
ligneous  cells  takes  up  from  48  to  51  per  cent,  while  the  jelly-like 
membranes  of  nostocacese  and  palmellacese  absorb  as  much  as  200 
per  cent  of  water,  according  to  NÄgeli;  on  the  other  hand,  cork 
membranes  have  hardly  any  swelling  capacity  at  all. 

The  resistance  offered  to  loss  of  water  is  exceptionally  variable. 
It  may  be  said,  with  certain  exceptions,  that  plants  are  much  more 
resistant  than  animals.  Especially  the  lower  forms  of  life,  particu- 
larly the  spores  of  bacteria,  yeasts,  algse,  mosses  and  seeds  may  bear 
almost  complete  dehydration  without  dying.  Loss  of  water  is  often 
of  great  biological  significance  for  plants.  It  makes  spores  and  seeds 
less  sensitive  to  changes  of  temperature;    and  in  the  case  of  some 

^  The  greater  water  content  of  the  individual  organs  of  the  newborn  as  con- 
trasted with  those  of  adults  is  especially  evident  from  the  tables  of  E.  Bischoff.* 

215 


216  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

higher  plants,  the  spreading  of  the  seed  pods  upon  drying  leads  to 
movements  which  serve  to  distribute  the  seeds.  Best  known  is  the 
"blooming,"  or  the  swelling  of  the  "Jericho-rose."  Higher  animals  on 
the  other  hand  are  very  sensitive  to  losses  of  water:  frogs,  according 
to  Kunde,  may  withstand  a  gradual  loss  of  water  up  to  30  per  cent, 
but,  if  they  are  rapidly  dried,  they  perish  when  the  loss  is  only  18 
per  cent.  In  the  latter  case,  there  is  evidently  no  time  for  an  equali- 
zation in  the  distribution  of  the  water.  Thirsting  human  beings  also 
show  great  losses  of  water,  though  of  course,  there  are  no  data  as  to 
the  lethal  point.  A.  Durig  informs  me  that,  after  a  forced  march 
in  hot  weather,  he  lost  5  kg.  of  water.  The  investigations  of  N. 
ZuNTZ  and  Schumberg  on  marching  soldiers,  as  well  as  those  of  N. 
ZuNTZ  on  mountain  climbers,  showed  that  exercising  men  lost  water. 
Roughly  measured,  the  water  ingested  after  forced  marches  does 
not  replace  the  water  lost.  We  may  say  here,  anticipating  some- 
what, what  animal  experiments  of  H.  Gerhartz*  show,  that  the  loss 
of  water  affected  primarily  the  musculature  and  then  the  fluids  cir- 
culating in  the  organs. 

Freezing  (gefrieren)^  has  an  effect  on  the  organism  similar  to  the 
withdrawal  of  water.  It  was  formerly  believed  that  the  formation  of 
ice  burst  the  cell  walls  or  tore  the  protoplasm,  and  the  damage  from 
freezing  was  ascribed  to  these  gross  mechanical  influences.  It  was 
shown  by  the  investigations  of  A.  E.  Nägeli,  W.  Sachs,  H.  Molisch, 
and  Müller-Thurgau  that  these  views  were  false,  that  usually 
there  was  no  formation  of  ice  in  the  cell,  but  that  the  ice  crystals 
grew  between  the  cells  in  the  intercellular  spaces.  P.  Matruchot 
and  MoLLiARD*  studied  plant  cells  and  found  that  the  phenomena 
observed  in  drying  or  plasmolysis  resembled  those  induced  by  freez- 
ing (erfrieren).  H.  W.  Fischer,*^  as  the  result  of  exhaustive  studies, 
reached  the  conclusion  that  the  damage  done  to  animals  and  plants 
by  freezing  them  (gefrieren)  was  analogous  to  the  partially  irre- 
versible changes  produced  in  gels  by  glaciation.  He  believes  that 
the  adsorption  of  electrolytes  in  particular  is  thus  affected  unfavor- 
ably. If  a  solution  of  potato  starch  is  frozen  and  then  thawed  out, 
the  electrolytes  may  entirely  dissolve  again  but  the  starch  has 
become  insoluble.  Frozen  leaves  present  an  analogous  condition  in 
that  the  chlorophyl  is  no  longer  retained.     In  this  valuable  work  he 

1  Freezing  (erfrieren)  and  glaciation  (gefrieren)  must  not  be  confused.  A  plant 
or  an  animal  freezes  if  life  processes  cease  by  reason  of  the  low  temperature. 
This  temperature  depends  upon  the  nature  of  the  organisms  involved,  and  in 
the  case  of  warm-blooded  individuals  is  usually  far  above  zero;  in  the  case  of 
other  organisms  (seeds  and  spores),  however,  it  may  be  far  below  zero  (—200). 
Glaciation  always  means  ice  formation. 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     217 


shows  that  the  condition  and  age  of  protoplasm  when  frozen,  i.e.,  at 
the  lethal  point,  is  similar  to  that  observed  by  van  Bemmelen  in  the 
drying  of  colloids;  when  the  latter  are  frozen,  they  become  optically 
inhomogeneous,  and  their  staining  capacity  changes. 

For  normal  functioning  there  is  an  indispensable  normal  water 
content  for  every  individual  or- 
ganism and  organ. 

The  total  water  content  of  an 
organism  gives  us  an  idea  of  its 
water  requirements;  the  water 
content  of  the  individual  organs 
informs  us  about  the  distribution 
of  water  in  the  organism. 

A  clearer  idea  is  obtained  by 
observing  the  distribution  of 
water  in  animals  especially  in 
mammals.  In  Table  III  (see  p. 
219),  I  have  compiled  the  water 
content  of  the  different  adult  organs 
from  available  data.  Table  II 
(p.  218)  shows  the  distribution 
of  the  total  water  content  (100 
per  cent)  in  the  various  organs. 
I  have  placed  the  proportionate 
weights  of  the  organs  to  the 
total  weight  alongside  for  com- 
parison. The  data  in  Tables  I 
and  II  show  far-reaching  differ- 
ences (see  especially  Skin  in 
Table  II),  for  which  age  and  nu- 
tritive condition  are  responsible. 

In  healthy  animals  and  men,  there  is  a  definite  swelling  ratio  for 
the  individual  organs,  a  dynamic  equilibrium.  The  maximal  varia- 
tion of  normal  swelling,  more  or  less,  is  called  the  swelling  range.^     It 

1  Definitions: 

Swelling  capacity  is  the  maximal  capacity  for  absorbing  water,  expressed  in 
W  (weight  of  water) 
D  (dry  weight) 

Swelling  is  the  water  content  of  a  gel  or  an  organ  expressed  in 

W 
D  ' 

Swelling  range  is  the  greatest  variation  in  the  capacity  of  an  organ  to  absorb 
water  under  different  conditions. 

•  W  (maximal  weight  of  water)  —  W  (minimal  weight  of  water) 

D  ■ 

A  normal  swelling  range  and  an  abnormal  swelling  range  exist. 


I 


a  6  c 

Fig.  37.  Spirogyra:  a,  before  the  ex- 
periment; b,  frozen;  c,  after  thawing. 
(From  H.  Molisch.) 


218 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


TABLE  II. 

Percentage  Distribution  of  Water  in  the  Individual  Organs. 

(Entire  water  in  the  organism  =  100  per  cent.) 

(After  A.  Albu  and  C.  Neuberg,*  Bischoff,*  Engels  *  and  A.  W.  Volkmann.*) 


Human. 

Dog. 

Distribution  of 

total  water  in 

each  organ. 

Weight  of  organs  in  per  cent 
of  body  weight. 

Distribu- 
tion of 
water  in 
each  organ. 

Weight  of 

otgans  in 

per  cent  of 

body 

weight. 

Blood     

4.7—9 
2.3 

6.6—11.0 

3.2 
2.8 
2.4 
0.4 
0.6 
9—12.5 

47.74—50.8 

2.7 

4.9 
Newborn             13.5 
Man                     18.2 
Woman               28.2 
Newborn            11.3 
Man                       6.9 
Woman                 5 . 7 

Man                      4.1 
Woman                 5 . 4 

8.27 

11.58 

9.68 
3.86 
2.83 

1.01 
9.08 

47.74 
1.59 

7 

Fat 

Skin 

16.11 

Viscera: 
Intestines 

8.18 

Liver 

3.60 

2.36 

Spleen 

Kidneys           

0.85 

Bony  skeleton 

Muscles 

Newborn  15.7-17.7 
Adult                  15.9 
Newborn  22.9-23.5 
Man                     41.8 
Woman               35.8 

Newborn  12.2-15.8 
Man                      2.6 
Woman                 2.7 

17.39 
42.84 

Nerve  substance: 
Brain 

Spinal  cord .           . .  . 

1.37 

Remainder 

11.0 

is  very  small  for  the  skeleton,  the  blood,  and  the  intestines,  has  a 
middle  value  for  the  viscera,  and  becomes  higher  for  skin,  muscles, 
and  kidneys. 

This  was  deduced  especially  from  the  experiments  of  Engels.* 
He  kept  dogs  without  food  four  days  and  determined  the  water 
content  of  various  organs  (normal  animals).  Another  series  of  dogs, 
after  the  same  preUminary  treatment,  received  an  infusion  into  their 
jugular  veins  of  1160  gm.  physiological  salt  solution  (on  the  average). 
Three  hours  after  the  termination  of  the  infusion,  the  animals  were 
killed  and  the  water  content  of  the  organs  determined  (water  treated 
animals). 

In  the  following  table,  the  percentage  of  the  water  infused,  that  is, 
recovered  from  the  individual  organs,  is  shown  in  column  A,  and  in 
column  B  is  the  percentage  increase  in  weight  of  the  several  organs 
in  terms  of  their  own  weight : 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     219 

(Basis  of  the  swelling  range.) 


A. 

ß. 

Muscles 

67.89 
17.75 
2.96 
2.25 
1.97 
1.55 
1.41 
1.13 
0.28 
2.82 

100.01 

17.1 

11.9 
8.9 
3.0 
9.0 
2.4 

17.9 
8.9 

10.0 

Skin 

Liver 

Intestine 

Lungs 

Blood 

Kidneys 

Brain 

Uterus 

Lost  in  bleeding 

TABLE  III. 

Water  Content  of  Various  Human  Organs  in  Per  Cent.^ 

(After  Albu-Neuberg,*  Bischoff,*  Halliburton,*  Pfibram,*i  Rumpf.*) 


Adult 
(normal). 

Child  (normal). 

N  —  Newborn. 

2  M  —  2  months  old. 

Adult  (pathological). 

Blood 

Fat 

77  9— 83 

29.9 
73.3—77 
79.2—80.2 
68.3—79.8 

78—79 
75.8—86.1 
77—83.7 
22—34 
73—75.7 

75—82 

N85 

N83.1 
2  M  75.5 
N  83.3—93.1 
2M80 
N  80.5 
2M73 
N82.6 
2  M  79.4 
N78.4 
2  M  77.7 
N  85.7 
2M81 
N32.3 
2  M  62.3 
N81.8 
2M71.7 

N89.3 
2M89 

90  and  more  (Anemia) 
75.5—83.9  (Nephritis) 
73.2—66.5  (Diabetes) 

Viscera: 

Intestines 

Heart 

79.2—80.4  (Nephritis) 
68.5—87.3  (Nephritis) 

Liver 

Lung 

Spleen 

90.6  (Nephritis) 
84.8—88.2  (Nephritis) 

• 

Kidneys 

Bony  skeleton 

Muscles 

Nerve  substance: 
Brain 

80.9—83  (Nephritis) 

1  I  have  omitted  the  figures  for  skin;  they  vary  for  different  authors  between  31.9  and  73.9,  be- 
cause some  have  given  the  water  content  of  skin  deprived  of  fat  and  others  that  with  the  fat  attached. 

Since,  in  the  dog,  muscles  constitute  42,82  per  cent  and  skin  16.11 
per  cent  of  the  total  body  weight,  these  organs  under  normal  con- 
ditions actually  store  up,  respectively,  47.74  per  cent  and  11.58  per 
cent  of  all  the  water  in  the  body.  As  a  result  of  their  great  swelling 
range,  tlie  two  chief  water  excreting  organs,  the  skin  and  kidneys,  the 


220  COLLOIDS  IN  BIOLOGY   AND  MEDICINE 

muscles,  most  of  all,  are  able  to  accumulate  large  quantities  of  water. 
In  Engel's  experiment  they  took  up  2/3  of  the  water  supplied. 

If  water  is  suppHed  to  the  organism,  the  blood,  on  account  of  its 
low  swelling  range,  gives  off  the  excess  chiefly  to  the  muscles  and 
skin;  it  parts  with  some  to  the  glands,  chiefly  the  kidneys.  On  this 
account,  saline  infusions  after  a  severe  loss  of  blood  have  usually 
only  a  temporary  effect.^  Muscles  and  skin  behave  like  a  reservoir, 
the  blood,  like  a  rigid  pipe  system,  from  which,  if  the  pressure  is 
sufficient,  excess  of  water  constantly  flows  through  a  small  vent.  In 
carrying  out  this  wise  arrangement,  the  organism  utilizes  the  various 
swelling  ranges  of  the  organ  colloids. 

Though  I  have  used  the  above  picture  of  a  rigid  pipe  system  for 
the  blood,  it  is  not  strictly  accurate,  for  the  blood  has  a  small 
swelling  range  of  its  own,  as  we  see  from  Engel's  table.  This  may 
be  attributed  to  the  fibrinogen,  as  we  learn  from  the  following 
facts.  The  required  data  I  have  taken  from  E.  Abderhalden 
(pp.  592-593). *i 

The  water  content  of  various  animals  is: 

Per  mil. 

'  In  the  entire  blood 749-824 

Serum 902-926 

Blood  corpuscles 604-633 

From  this  we  see  that  when  the  water  content  of  serum  increases 
2.6  per  cent  (from  the  minimum),  it  reaches  its  maximum,  and  the 
blood  corpuscles  reach  their  maximum  with  an  increase  of  5  per 
cent.  The  entire  blood  on  the  contrary  has  a  swelling  range  of  10 
per  cent.  There  must  therefore  be  something  in  the  blood  that 
swells  especially  well  and  the  only  possible  substance  is  the  fibrinogen. 

Let  us  compare  the  maximal  and  minimal  content  of  water  dis- 
tributed between  serum,  blood  corpuscles,  and  the  whole  blood  in  the 
identical  animals. 

Max.  =  maximum  water  content  among  the  various  species  of 
animals. 

Min.  =  minimum  water  content  among  the  various  species  of 
animals. 

^  Attempts  to  hold  the  fluid  in  the  vessels  by  the  addition  of  colloids  have 
been  unsatisfactory.  A  more  favorable  result  is  obtained  when  14  grams  of  salt 
and  10  grains  of  crystalline  sodium  carbonate  are  administered  either  intrave- 
nously or  by  rectum  (J.  J.  Hogan  and  M.  H.  Fischer).  It  was  accomplished 
through  reducing  the  swelling  of  the  other  tissues  by  hypertonic  saline  and  neu- 
tralization of  acid  by  the  alkali.  Cholera  collapse,  which  results  from  the  water 
deprivation  by  reason  of  diarrhoea,  may  be  successfully  combated  by  hypertonic 
saline  infusions  (Roger)-.  [W.  M.  Bayliss  and  M.  H.  Fischer  have  recommended 
the  use  of  gum  arable  solution,  and  it  is  being  successfully  employed  at  the  front 
in  the  present  war,  see  p.  137.    Tr  j 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL      221 


Serum 

Corpuscles. 

Entire  blood. 

Cat 

Horse  I 

Sheep  I  

926Voo  (max.) 
902Voo(min.) 
917Voo 
925Voo 

624Voo 
613Voo 

604Voo(min.) 
633Voo  (max.) 

795Voo 

749Voo(min.) 

821Voo 

Rabbit 

817Voo 

The  serum  of  the  cat  had  a  higher  water  content  than  that  of  the 
other  animals  experimented  upon;  the  corpuscles  also  had  a  high 
water  content,  even  though  not  the  highest;  the  entire  blood,  how- 
ever, is  only  a  little  above  the  average.  For  Horse  I,  serum  and 
entire  blood  have  minimum  values  and  the  corpuscles  a  low  but 
by  no  means  the  lowest  value.  In  the  case  of  Sheep  I,  the  entire 
blood  reaches  a  maximum,  whereas  the  blood  corpuscles  show  a 
minimum,  and  the  serum  possesses  a  water  content  a  little  above 
the  average.  With  the  rabbit,  there  is  a  high  water  content  in  all 
portions.  From  this  we  learn  that  a  substance  possessing  a  great 
range  of  swelling  exists  in  the  whole  blood,  namely,  the  fibrinogen. 
We  recognize  further,  that  the  elements  of  the  blood  possess  a  cer- 
tain elasticity,  which  smoothes  out  the  fluctuations  in  the  water 
content,  and  which  shows  itself  by  a  "  Zog  "  of  the  water  plenitude 
or  poverty  in  the  various  elements  of  the  blood,  depending  on 
whether  there  is  a  supply  or  a  withdrawal  of  water,  a  swelling  or  a 
shrinking. 

The  following  swelling  ranges  are  found  for  the  various  elements 
of  the  blood:  fibrin  >  whole  blood  >  corpuscles  >  serum.  It  would 
be  desirable  to  have  investigations  of  the  water  content  of  the  differ- 
ent elements  of  the  blood  in  the  same  animal  before  and  after  water 
has  been  given. 

What  is  it  then,  that  determines  the  water  content  or  swelling  of  an 
organ?  Undoubtedly  each  organ  colloid  has  a  definite  swelling  ca- 
pacity and  a  definite  swelling  range.  A  priori,  we  may  assume  that 
the  colloids  of  muscles  swell  more  than  those  of  the  epidermis. 
Without  doubt,  the  structure  of  the  given  colloid  is  also  a  factor. 
W.  Pfeffer*^  {loc.  cit.  I,  p.  61)  justly  emphasizes  the  distinction 
between  water  of  swelling,  consequent  upon  the  hydrophile  state  of 
the  swelling  substances  and  the  water  of  imbibition,  which  is  drawn 
up  into  the  capillary  interstices  as  into  a  sponge.^ 

E.  Pribram*^  beUeves  that  the  sweUing  of  protoplasm  in  its  true 
sense,  i.e  ,  of  the  assimilated  (species-native)  colloids  of  the  cell  is  con- 

1  W.  Pfeffer  speaks,  it  is  true,  of  "molecular"  water  of  imbibition  (or  ad- 
herent water)  and  of  "capillary"  water  of  imbibition,  yet  he  intends  the  same 
distinction  that  I  have  indicated  above. 


222  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

stant.  Only  the  swelling  of  the  nonassimilated  reserve  substance  is 
variable.  This  view,  it  seems  to  me,  receives  its  chief  support  from 
the  findings  of  H.  W.  Fischer  and  P.  Jensen*  on  muscle,  which  is 
treated  more  thoroughly  on  page  291.  According  to  their  findings, 
water  occurs  in  muscle  in  two  phases.  One  has  a  constant  value 
and  is  closely  associated  with  the  viability  of  muscle  (conditioned  by 
the  integral  muscle  protoplasm).  The  other  phase  varies  in  the  ex- 
ercising muscle  and  is  only  loosely  bound  (water  of  swelling  of  the 
reserve  material). 

Besides  these  factors  of  swelling  which  are  inherent  in  the  organ 
colloids  under  consideration,  there  are  others  which  are  to  a  certain 
extent  impressed  from  without.  The  natural  salt  content  as  well  as 
the  products  of  metabolism,  especially  the  acids,  are  determinative  of 
the  swelling  of  a  tissue.  Acid  formation  in  an  organ  {e.g.,  CO2  or  lactic 
acid  in  active  muscle;  CO2  in  blood  corpuscles)  increases  its  swell- 
ing capacity.  If  we  observe  that  the  potassium  salts  predominate  in 
one  organ,  and  in  others,  the  sodium  salts,  or  even  that  there  is  an 
accumulation  of  salts  in  a  certain  portion  of  a  single  cell,  we  may 
conclude  from  that  alone,  that  the  water  content  also  depends  upon 
such  concentration.  Potassium  salts  increase  swelling;  Ca  salts 
deplete  (E.  Widmark*),  and  according  to  R-.  Chiari  and  Januschke 
inhibit  exudation. 

When  the  loss  of  water  is  very  great  (cholera,  infant  diarrhoeas), 
there  is  an  increase  of  potassium  salts  and  phosphates  in  the  urine. 
From  this  it  may  be  assumed  that  Na  salts  replace  the  K  salts  of 
the  muscles,  and  at  the  same  time  water  is  given  off.  According  to 
E.  Pribram, *2  this  occurs  in  order  to  protect  more  vital  organs,  espe- 
cially the  brain,  from  loss  of  water. 

Little  is  known  concerning  swelling  from  a  biological  point  of 
view.  On  this  account,  an  observation  of  H.  Paul*  seems  especially 
noteworthy.  He  pointed  out  in  the  case  of  peat  mosses  that  the  high 
moor  sphagnum  is  able  to  absorb  much  more  water  than  the  low  moor 
sphagnum.  For  example,  sphagnum  molluscum  absorbs  27  times,  and 
sphagnum  platyphyllum  absorbs  16  times  its  dry  weight  of  water. 
In  the  same  paper  we  find  that  the  high  moor  sphagnum  contains 
much  more  acid  than  the  low  moor  sphagnum,  and  that  the  former 
are  much  more  sensitive  than  the  latter  to  the  action  of  alkalis,  lime 
and  salt.  From  this  it  seems  to  me  we  may  certainly  infer  that 
swelhng  of  high  moor  sphagnum  is  greater  than  that  of  low  moor 
sphagnum  because  of  its  greater  acidity,  and  that  the  damage  it 
suffers  from  salts,  etc.,  may  be  attributed  to  the  alteration  in  its 
normal  condition  of  swelling. 

We  shall  see  that  abnormal  accumulation  of  acid  in  the  tissues 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     223 

results  in  their  swelling  or  edema.  We  recognize  from  this,  that  the 
dynamic  balance  of  the  swelling  is  dependent  upon  the  normal 
course  of  the  processes  of  assimilation  and  dissimilation.  Con- 
versely, pathological  processes  are  always  followed  by  an  abnormal 
condition  of  swelling. 

Pathology  of  Water  Distribution. 

In  pathological  conditions,  the  water  content  may  have  values 
very  different  from  normal.  The  water  in  the  blood  rises  to  90  per 
cent  and  more  in  severe  anemias,  and  may  fall  to  from  73.2  to  66.5 
per  cent  in  diabetes  (see  p.  219).  Under  other  pathological  condi- 
tions, organs  may  show  abnormal  swelling  (see  last  column  in  Table 
III,  p.  219). 

With  the  active  metabolism  and  formation  of  crystalloids,  which 
occur  in  fever,  there  are  alterations  in  swelling  (thirst,  dryness  of 
the  skin),  the  exact  nature  of  which  we  do  not  as  yet  understand. 

As  a  rule,  there  has  been  less  attention  paid  to  the  study  of  the 
conditions  in  which  the  swelling  of  an  organ  is  below  normal.  The 
injection  of  protoplasmic  poisons  (some  heavy  metal  salts,  strong 
acids)  causes  a  coagulation  of  the  organ  albumen,  which  reduces  to  a 
greater  or  less  extent  its  swelling  capacity.  I  am  still  occupied  with 
more  exhaustive  studies  of  these  questions  which  are  also  touched 
upon  in  the  chapter  on  Necrosis.  It  is  too  early  to  report  the 
results. 

Edema. 

By  edema  we  understand  an  abnormal  collection  of  fluid  in  tissue 
or  tissue  spaces;  if  the  fluid  collects  abnormally  in  a  body  cavity,  we 
call  it  an  exudate  or  hydrops. 

The  view  most  generally  accepted  up  to  a  few  years  ago  was  that 
edema  occurred  whenever  the  venous  blood  pressure,  or  more  cor- 
rectly, the  difference  between  arterial  and  venous  pressure  was  gen- 
erally or  locally  raised,  and  the  resistance  of  the  vessel  walls  was 
diminished  (Julius  Cohnheim,  1877).  It  is  known  that  in  heart 
disease  and  in  nephritis,  when  the  circulation  is  disturbed,  that  large 
portions  of  the  body,  especially  the  lower  extremities,  swell  and 
become  edematous.  The  local  inflammatory  edema  accompanying 
inflammation;;  insect;bites  or  the  injection  of  an  irritating  fluid  (e.g., 
diphtheria  toxin)  must  also  be  considered. 

The  above  explanation-  has  some  fascination,  and  it  cannot  be 
denied  that  it  will  continue  to  be  invoked  in  explanation  of  certain 
points,  especially  since  the  observation  of  increase  in  the  permeability 
of  vessel  walls  cannot  be  avoided,  when  we  see  that  even  corpuscles 


224 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


I'iG.  38.    The  ligated  leg  (right)  of  the  frog  is  edematous. 
(From  M.  H.  Fischer.) 

Reproduced  from  Fischer's  "  Oedema  and  Nephritis,"  second  edition,  by  permission  of  Messrs. 
John  Wiley  &  Sons,  Inc. 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     225 

pass  through  the  walls.  In  general,  however,  it  may  be  asserted 
that  the  above  explanation  has  brought  no  advance  to  our  under- 
standing of  edema.  Histologists  and  physiologists  have  contributed 
an  interminable  amount  of  work  without  making  any  progress. 

A  fundamental  departure  in  the  understanding  of  edema  and  its 
associated  questions  was  made  in  1907  by  the  investigations  of  an 
American,  Martin  H.  Fischer^,  who  sought  the  cause  of  edema,  not 
in  the  vessels,  but  in  the  tissues  themselves;  he  attributes  to  the 
edematous  tissues  an  increased  swelling  capacity.  M.  H.  Fischer's 
views  were  immediately  contradicted  and  led  to  more  experimental 
studies  and  scientific  discussions  than  most  biological  theories. 
However,  though  we  must  now  admit  that  M.  H.  Fischer's 
views  are  too  far  reaching,  it  cannot  be  denied  that  he  set  up  a  very 
productive  working  hypothesis.  We  shall  first  state  his  theory  and 
then  discuss  his  opponents'  views. 

The  most  important  experiment  of  M.  H.  Fischer  is  the  follow- 
ing :  he  ligatures  the  hind  limb  of  a  frog  so  that  its  circulation  is  cut 


Fig.  39.     Rabbit's  kidneys;    left  normal,  right  experimentally  edematous. 

off  (Fig.  38)  and  places  it  in  water  so  that  the  limbs  are  covered. 
The  hgatured  limb  swells  up  and  at  the  end  of  2  or  3  days  may  be 
2  or  3  times  its  original  weight.  If  the  frog  is  kept  in  a  dry  vessel, 
the  ligatured  limb  dries  up  completely,  and  if  it  is  cut  off  and  placed 
in  water,  it  swells  up.  Under  these  conditions  the  blood  pressure  or 
the  increased  permeability  of  the  vessel  walls  cannot  play  any  part 
in  the  development  of  the  edema,  but  it  is  only  the  tissues,  which  swell 
more  strongly  under  the  circumstances  mentioned.  In  the  same 
manner,  M.  H.  Fischer  was  able  to  demonstrate  the  occurrence  of 
edema  in  rabbits'  kidneys  (Fig.  39),  and  in  the  livers  and  lungs  of 

^  He  was  led  to  this  by  experiments  of  Jacques  Loeb,  which  showed  that 
frogs  muscles  swelled  more  in  acid  and  alkaline  fluids  than  in  neutral  ones. 


226  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

f 
sheep.    These  became  edematous,  not  only  if  the  veins  were  hgatured, 

but  also  if  the  artery  was  tied.  Under  the  circumstances,  an  in- 
creased blood  pressure  as  the  result  of  congestion  need  not  be  con- 
sidered. Indeed,  all  dead  bodies  or  portions  of  them  which  certainly 
are  without  blood  pressure  swell  up  when  immersed  in  water. 

It  was  thus  demonstrated  that  the  development  of  edema  de- 
pended upon  the  increased  capacity  of  the  tissues  to  swell,  and  the 
question  then  presented  itself,  What  change  in  the  tissues  permits  the 
development  of  edema? 

An  explanation  was  provided  by  the  studies  of  F,  Hofmeister,  his 
pupils  and  successors,  upon  the  swelling  capacity  of  gelatin  and  re- 
lated substances  (see  pp.  67-68).  We  learned  in  that  chapter  that 
acids  and  alkahes  increase  swelling  capacity,  that  other  electrolytes 
in  the  order  there  mentioned,  either  favor  or  diminish  swelling,  while 
nonelectrolytes,  as  far  as  appropriate  studies  as  yet  show,  have  only 
slight  influence.  Martin  H.  Fischer*  (in  collaboration  with 
Gertrude  Moore)  was  able  to  demonstrate  that  the  same  laws 
governed  the  swelling  capacity  of  fibrin,  the  swelling  of  frogs' 
muscles  and  the  extirpated  eyes  of  oxen  and  sheep.  This  explana- 
tion presupposes  no  membrane  or  osmotic  pressure;  it  permits  an 
unstrained  interpretation  of  processes  which  otherwise  are  explained 
with  great  difficulty  in  the  case  of  animal  cells  unprovided  with 
membranes. 

The  further  question  now  presents  itself.  What  electrolytes  are  re- 
sponsible for  the  altered  swelling  capacity  of  the  tissues  in  edema? 
We  can  no  longer  offer  a  single  explanation,  and  we  must  study  indi- 
vidual cases. 

Edema  fluid,  the  CO2  of  which  has  been  removed,  is  acid  to 
Phenolphthalein;  F.  Hoppe-Seyler  found  in  it  valeric,  succinic, 
butyric  and  lactic  acids.  Strassburg  and  R.  Ewald  found  that  the 
CO2  content  of  edema  fluid  was  greatly  in  excess  of  that  of  venous 
blood. ^  Of  especial  value  is  the  discovery  by  F.  Araki  and  H. 
ZiLLESSEN  that  any  lack  of  oxygen  is  followed  by  an  excessive 
production  of  acid,  though  this  fact  may  not  be  demonstrable  by 
indicators.  ' 

One  answer  is  thus  given  to  the  question  asked  above.  Increased 
acid  production,  one  of  the  results  of  deficient  oxygen,  may  cause  the 
development  of  edema.  Such  a  condition  occurs  in  circulatory  dis- 
turbances and  in  cardiac  insufficiency,  where  edema  is  especially  fre- 

1  I  wish  to  point  out  that  a  certain  contradiction  is  contained  in  the  simul- 
taneous presence  of  organic  acids  and  increased  CO2.  Possibly  this  may  be 
attributed  to  the  fact  that  various  edema  fluids  have  been  examined  and  un- 
justified generaUzations  deduced.     (The  author.) 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL      227 

quent;  it  occurs  also  in  severe  anemias  and  in  certain  cachexias, 
starvation  and  scurvy.  In  nephritics,  some  substances  recently  dis- 
covered in  the  blood,  may  perhaps  contribute  to  the  inhibition  of 
oxidation.  Cadaveric  edema  is  well  known,  as  is  the  bloated  ap- 
pearance of  drowned  bodies.  In  the  case  of  living  animals,  only  the 
injection  of  excessive  quantities  of  water  or  physiological  salt  solu- 
tion are  able  to  bring  about  edema.  According  to  R.  Magnus,  this 
is  readily  accomphshed  by  injecting  salt  solution  into  the  blood 
vessels  of  dead  animals.  A  dead  frog  may  double  its  weight  in  from 
36  to  48  hours  if  immersed  in  water. 

The  injection  of  certain  poisons  (especially  lactic  acid)  results  in 
an  oxygen  deficiency  and  thus  brings  about  an  excessive  acid  pro- 
duction. M.  H.  Fischer  injected  morphin,  strychnin,  cocain, 
arsenic  and  uranium  nitrate  into  the  dorsal  lymph  sac  of  frogs  and 
thus  produced  an  edema  which  disappeared  if  the  frog  was  given  an 
opportunity  to  excrete  the  poison. 

Edema  in  the  case  of  metal  intoxications,  especially  in  the  case  of 
arsenic,  is  well  known  to  clinicians,  and  the  great  thirst  and  the  dimi- 
nution in  the  excretion  of  urine  which  occurs  after  morphin,  ether  and 
chloroform  administration,  may  also  be  attributed  to  the  deficiency 
of  oxygen  in  the  tissues,  with  concomitant  absorption  of  water. 

We  have  only  referred  to  the  development  of  edema  by  acids,  and 
I  wish  to  call  attention  to  the  fact  that  intense  edema  may  be  pro- 
duced by  alkalies.  Subcutaneous  injection  of  n/10  sodium  hydrate 
results  in  severe  edema. 

If  such  substances  produce  edema  as  favor  the  swelling  of  gelatin, 
fibrin,  etc.,  edema  must  be  counteracted  by  electrolytes  which  re- 
duce swelling.  The  correctness  of  this  assumption  was  demonstrated 
by  M.  H.  Fischer  on  the  amputated  leg  of  a  frog.  The  addition  of 
neutral  salts  diminished  the  swelling  and  acted  in  the  same  order 
that  the  cations  and  anions  did  in  diminishing  the  swelling  of  fibrin. 
Nonelectrolytes,  on  the  other  hand,  had  no  influence. 

M.  H.  Fischer  regards  glaucoma  as  a  typical  example  of  a  local 
edema.  This  is  a  disease  of  the  eyes,  of  which  the  most  character- 
istic symptom  is  very  greatly  increased  tension,  which  produces 
hardness  of  the  eyeball.  The  excruciating  pains  and  loss  of  vision 
are  mere  consequences.  The  various  explanations  given  in  ophthal- 
mological  textbooks  are  quite  unsatisfactory,  whereas  the  experi- 
ments of  M.  H.  Fischer  are  quite  convincing.  M.  H.  Fischer 
placed  extirpated  ox  eyes  ^  in  water,  to  which  so  little  acid  had  been 
added  that  it  was  imperceptible  to  taste.     These  eyes  became  stony 

1  The  investigations  of  P.  Bottazzi*  and  his  pupils  on  the  sweUing  and 
shrinldng  of  lenses  in  solutions  of  acids,  bases  and  salts  should  be  mentioned. 


228  COLLOIDS  IN  BIOLOGY  AND   MEDICINE 

hard  as  in  the  most  aggravated  glaucoma.  On  the  other  hand,  by  the 
injection  of  a  few  drops  of  1/8  to  1/6  molecular  (4,05  to  5.41  per  cent) 
sodium  citrate  solution  under  the  conjunctiva,  it  was  possible  to  cause 
the  pressure  in  human  glaucoma  and  in  artificially  glaucomatous  eyes 
to  become  normal  or  even  subnormal  in  less  than  five  minutes.  [The 
observation  of  Riesman  on  the  lowered  ocular  tension  of  diabetics  and 
the  lowering  of  ocular  tension  by  intravenous  injection  of  glucose  by 
Woodyatt's^  method  are  important  in  this  connection.     Tr.] 

Cloudiness  of  the  cornea  occurs  without  reference  to  the  absorption 
of  water  by  the  eye.  It  probably  depends  on  the  precipitation  of  a 
protein,  since  all  acids,  bases,  salts  and  nonelectrolytes  which  cause  a 
precipitation  of  proteins  cause  a  corneal  turbidity.  What  is  true  of  the 
cornea  may  be  supposed  to  apply  also  to  the  other  transparent  media 
of  the  eye,  the  lens  and  the  vitreous  humor.  Here,  too,  therapeutic 
results  have  been  obtained  already,  by  injecting  sodium  citrate. 

Hayward  G.  TnoivLiS  obtained  an  improvement  in  vision  in  cases 
showing  a  cloudiness  of  the  cornea,  the  lens  or  the  vitreous  humor. 
It  is  natural  to  assume  that  the  turbidity  is  due  to  a  reversible  pre- 
cipitation of  albumin.  Similar  results  were  obtained  in  an  edema  of 
the  tissues  surrounding  the  knee. 

The  tiny  sweUings  which  result  from  insect  bites  are  regarded  by 
M.  H.  Fischer  as  local  edemas  produced  by  a  drop  of  acid  or  some 
substance  which  interferes  with  the  normal  oxidative  processes  of 
the  tissues.  The  beneficial  action  of  ammonia,  customarily  employed 
on  insect  bites,  favors  this  view.  M.  H.  Fischer  produced  "artificial 
flea  bites  "  on  gelatin  plates  by  sticking  them  with  needles  dipped  in 
formic  acid  and  then  placing  them  in  water;  with  ammonia  he  was 
able  to  make  the  swellings  recede. 

One  of  M.  H.  Fischer's  observations  seems  of  great  significance  to 
me  in  explaining  the  phenomena  of  some  skin  diseases.  He  ob- 
served in  gelatin  plates  upon  which  moulds  had  been  sown,  an  ele- 
vation in  the  centre  of  each  colony  (swelling).  If  we  recall  that  many 
skin  diseases  are  caused  by  true  fungi  related  to  the  moulds,  and 
that  in  many,  the  most  characteristic  symptoms  are  wheals  (swell- 
ings), papules  and  vesicles,  the  analogy  cannot  be  neglected.  More- 
over, we  must  think  of  analogous  processes  if  we  consider  other  local 
edemas  occurring  after  inoculation  with  infectious  microorganisms 
or  the  injection  of  diphtheria  antitoxin. 

In  a  later  work,  M.  H.  Fischer*^  elaborated  his  fundamental 
ideas  of  cellular  pathology  and  studied  cloudy  swelling.  Cloudy 
swelling  is  found  in  the  liver,  kidneys,  spleen,  and  muscle  cells  of 

1  Studies  in  Intermediate  Carbohydrate  Metabolism.  R.  T.  Woodyatt. 
Harvey  Lecture,  1915-1916,  p.  328. 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     229 

persons  dying  as  the  result  of  acute  infectious  diseases.  The  cells 
involved  are  enlarged  and  more  or  less  cloudy;  macroscopically,  the 
organ  involved  appears  at  times  as  if  it  had  been  boiled;  microscopic- 
ally, granular  deposits  are  seen. 


steamy 


(yellowish') -jviilte 
white  aucleua 


Fig.  40. 

Reproduced  from  Fischer's  "Oedema  and  Nephritis,"  second  edition,  by  permission  of  Messrs. 
John  Wiley  &  Sons,  Inc. 

M.  H.  Fischer  experimentally  produced  cloudy  swelling  in  the 
livers  and  kidneys  of  rabbits  by  placing  them  in  distilled  water  or 
dilute  acids.  Addition  of  various  salts  delayed  or  hastened  the  de- 
velopment of  cloudy  swelling.  According  to  M.  H.  Fischer,  whether 
the  swelling  occurs  pathologically  or  experimentally,  it  is  explained 
just  as  is  edema;  as  the  result  of  the  production  of  acids  in  the 
injured  tissues,  they  swell.  The  cloudiness  runs  parallel  with  the 
precipitation  of  proteins,  especially  that  of  casein  by  acids.  The  pro- 
duction and  disappearance  of  granules  in  parenchymatous  cells  in 
the  presence  of  additional  acid  occurs  exactly  hke  the  precipitation 
a,nd  re-solution  of  casein  upon  increasing  the  concentration  of  acid. 
The  "cloudy  swelling"  is,  therefore,  the  result  of  acid  production  in 
the  tissues,  though  the  swelling  and  the  clouding  are  two  processes 
entirely  independent  of  each  other. 

As  has  been  stated  already,  M.  H.  Fischer's  theory  was  actively 
discussed  and  contradicted.  Before  approaching  this  discussion  we 
shall  present  a  brief  resume.     Fischer  maintains  that  edema  results 


230  COLLOIDS  IN  BIOLOGY  AND   MEDICINE 

from  the  swelling  of  organ  colloids,  which  is  induced  by  acids  that  are 
produced  by  a  disturbance  in  the  oxidative  processes  of  an  organ. 

The  first  objection  is  directed  against  the  assumption  that  processes 
which  occur  in  dead  colloids  may  be  transferred  to  the  living  or- 
ganism. It  was  raised  by  G.  Bentner,  Jacques  Loeb  and  A.  R. 
Moore.  They  admit  that  Fischer's  experiments  apply  to  dead 
tissue,  especially  muscle,  but  that  they  fail  in  the  living  organ  where 
osmotic  processes  are  active  and  satisfy  all  the  conditions.  The  objec- 
tion of  R.  Holer  is  especially  searching,  that  if  only  electrolytes  may 
inhibit  acid  swelling,  a  piece  of  muscle  would  swell  in  sugar  solution. 
As  a  matter  of  fact  the  muscle  volume  is  unchanged  in  an  isotonic  sugar 
solution.  M.  H.  Fischer  *^  combats  this  by  stating  that  the  existence 
of  osmotic  membranes  in  living  cells  is  quite  readily  conceivable  (see 
p.  290),  and  points  to  his  experiments  which  reproduced  the  contraction 
of  living  muscle  by  means  of  catgut,  a  dead  colloid  material.  Osmotic 
attraction  of  water  is  inconceivable  nor  does  it  assist  the  explanation. 

The  second  objection  is  directed  against  the  very  development  of 
acids  in  tissues  and  was  raised  by  A.  R.  Moore,  who  was  unable  to 
detect  acids  by  acid  fuchsin  or  neutral  red  in  muscles  made  edematous 
according  to  Fischer's  technic  nor  in  the  lymph  or  kidneys  of  rabbits 
injected  with  acid  salts;  and  who  maintained  that  consequently  the 
acid  content  of  such  tissues  is  not  responsible  for  the  edema  or  albu- 
minuria. Fischer  meets  this  objection  by  stating  that  acidity  of  tis- 
sues is  not  to  be  detected  by  color  indicators  since  the  acids  combine 
with  proteins  and  that  even  traces  of  acids  induce  swelling  and  that 
there  is  no  more  delicate  indicator  of  acidity  than  the  swelHng  of 
proteins.  [Fischer  insists  that  p^  swelling  in  protein  nowhere  par- 
allels Pb  concentration.  The  degree  of  swelling  follows  the  order, 
HCl  >  lactic  >  sulphuric  acid.     Tr.] 

The  third  objection  is  directed  against  the  generaHzation  of  Fisch- 
er's hypothesis.  Fischer  performed  his  experiments  principally  on 
muscles  which  behaved  by  swelling  in  the  presence  of  acids  Uke  fibrin 
or  other  dead  colloid  but  this  would  not  apply  to  all  kinds  of  tissue. 
Connective  tissue  and  cartilage  apparently  behave  the  same  way. 
L.  PiNCUSSOHN,  however,  found  that  kidney,  spleen,  liver  and  lung 
usually  became  less  swollen  in  acid  than  in  pure  water.  Kidney  cortex 
and  kidney  medulla  showed  a  difference  in  that  the  former  became 
more  swollen  in  acid  and  water  than  the  latter.  These  experiments  do 
not  impress  me  as  decisive  because  physiological  salts  were  absent. 

The  behavior  of  nerve  tissue  is  especially  interesting.  Reichardt 
called  attention  to  a  clinical  condition  which  occasionally  occurs  in 
dementia  prsecox  and  causes  sudden  death.  He  noted  an  increase  of 
weight  and  volume  in  such  brains  without  other  detectable  macro-  or 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     231 

microscopic  changes.  He  designated  this  condition  swelling  of  the  brain 
in  contrast  with  edema  of  the  brain;  it  is  possible  to  demonstrate  the  fluid 
transudate  from  the  blood.  In  swelHng  of  the  brain,  it  is  dry,  sohd  and 
gelatinous.  Reichardt's  observation  was  made  about  the  time  Fischer 
published  his  theory  so  that  it  seems  apropos  to  mention  it.  J.  Bauer 
and  Bauer  and  Ames  tested  slices  of  brain  and  cord  and  found  that 
swelling  was  observed  occasionally  with  a  one  thousandth  normal  acid- 
ity, but  with  greater  concentration  (even  of  carbonic  acid),  a  shrinking 
always  occurred.     On  this  account  they  reject  Fischer's  acid  theory. 

Contradicting  J.  Bauer,  Fischer  in  experiments  (with  Hooker) 
found  that  nervous  tissue  behaved  toward  salts  and  acids  Hke  fibrin 
and  other  tissue.  He  explains  the  disagreement  by  the  fact  that 
Bauer  chose  tissue  which  had  been  dead  6  to  24  hours.  The  optimal 
concentration  for  sweUing  had  already  been  exceeded  by  the  post 
mortem  development  of  acid. 

The  fourth  objection  was  raised  by  pathologists  who  maintained 
that  what  Fischer  described  was  not  edema  at  all.  The  main  loca- 
tion of  edema,  connective  tissue,  exhibits  apparently,  Hke  fibrin,  the 
swelfing  and  shrinking  phenomena  with  acids  and  salts  but  is  essen- 
tially dissimilar  from  edematous  tissue  and  more  like  hyaline  or  amy- 
loid degeneration.  (Marchand,  Klemensiewicz,  H.  Schade.)  The 
fibers  show  the  chief  swelhng  in  acids  but  in  edema  the  main  swelfing 
is  extrafibrillar.  Fischer  fails  to  distinguish  between  swelling  of  the 
protoplasmic  substance  and  turgor  of  the  entire  tissue.  Against  this 
view  that  in  edema  the  aqueous  fluids  accumulate  in  the  tissue 
spaces  and  not  merely  in  the  protoplasma  and  that  increased  inhibi- 
tion of  water  by  the  cell  is  not  the  criterion  of  edema,  M.  H.  Fischer 
argues  that  the  tissue  spaces  are  not  filled  with  air  but  by  colloid  ma- 
terial which  may  very  well  contribute  to  the  edema  by  acid  swelling. 

Lubarsch  noted  marked  swelfing  of  the  tissue  in  his  histologi- 
cal studies  but  determined  a  difference  in  kidney  edemas  due  to 
clamping  the  renal  artery  or  renal  vein.  The  changes  are  snxdlar; 
those  produced  by  clamping  the  vein  are  reversible  but  those  due  to 
clamping  the  artery  are  irreversible  if  the  artery  is  clamped  off  for 
three  hours.  The  sensitive  ceHs  are  kified.  This  contradicts 
Fischer's  theory  which  requires  the  damage  to  be  the  same  in  either 
case.  Kurt  Ziegler  made  the  üiiportant  observation  that  chlorid 
metabofism  as  weU  as  water  metabofism  play  an  important  part  in 
edema.  Chlorid  and  water  retention  alternate  as  primary  factors 
in  edema,  but  in  all  cases,  there  are  nutritive  disturbances  which 
affect  chiefly  muscles  and  connective  tissue.  P.  Tachau  offered 
experimental  verification  by  feeding  mice  excessive  amounts  of  sodium 
salts.     There  occurred  edema  about  the  head,  neck  and  attachment 


232  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

of  front  limbs  resembling  that  observed  in  human  nurslings  when 
they  are  given  too  much  salt.  In  Tachau's  experiments  it  is  note- 
worthy that  there  was  no  increase  in  average  amount  of  water  in  the 
animal  but  the  edema  was  the  expression  of  abnormal  distribution  of 
water.  These  observations  lend  some  support  to  Fischer's  theory. 
Fischer  observed  that  when  fibrhi  and  gelatin  swelled  up  they 
accumulated  salt  from  an  acid  table  salt  solution  and,  consequently, 
he  regarded  the  salt  retention  as  resulting  from  the  edema. 

When  we  consider  the  controversy  concerning  Fischer's  theory  of 
edema,  it  is  evident  that  as  yet  he  has  offered  no  experimental  proof 
for  his  hypothesis.  Except  in  a  few  special  instances  his  opponents 
have  likewise  failed  to  show  its  invalidity  since,  in  my  opinion,  their 
experimental  methods  failed  to  produce  a  local  accumulation  of 
acid  in  hving  organs  as  required  by  Fischer's  theory.  No  matter 
what  value  may  be  set  on  Fischer's  theory  in  the  future,  it  has  been 
of  enduring  service  in  that  it  has  transferred  the  emphasis  in  the  study 
of  edema  from  the  circulation  to  the  tissues;  it  is  not  hydrostatic 
differences  in  pressure  but  chemical  damage  to  the  tissue  which 
occasions  edema.^ 

Inflammation. 

Though  healthy  cells  are  impermeable  for  blood  plasma,  inflamed 
tissue  permits  a  selective  passage  of  plasma  elements. 

A.  Oswald  *  found  that  the  frequency  of  passage  stood  in  the  fol- 
lowing order: 
Albumin  >  Globulin  (Euglobulin)  >  Pseudoglobulin  >  Fibrinogen. 

This  occurs  in  an  order  inversely  to  their  susceptibility  to  salting  out 
by  salts  and  to  the  viscosity  of  the  various  solutions :  the  less  viscous 
a  plasma  element  is,  the  more  easily  does  it  pass  through  the  inflamed 
tissue.  Albumin  alone  may  be  found  in  an  exudate,  but  never  fibrin- 
ogen without  the  simultaneous  presence  of  albumin  and  globulin. 
In  the  acute  stages  all  kinds  of  albumins  are  found  in  the  exudate, 
whereas,  with  the  lapse  of  time,  fibrinogen  and  then  globulins  diminish. 
The  normal  cell  membrane  evidently  behaves  Hke  an  impermeable 
ultrafilter,  which  has  become  more  permeable  by  reason  of  the  in- 
flammatory process.  We  do  not  know  the  factors  which  bring  this 
about.  [The  tissue  may  be  ''coagulated"  allowing  freer  diffusion 
because  of  larger  diffusion  paths,  or  they  may  be  more  dispersed  and 
accumulate  more  "water  of  swelling."  Either  condition  would 
explain  some  of  the  phenomena.     Tr.] 

^  [The  most  recent  discussion  of  the  question  by  Lawrence  J.  Hendeeson  and 
Martin  H.  Fischer  is  contained  in  the  Journal  of  the  Am.  Chem.  Society, 
Vol.' XL,  No.  5  (May,  1918).    Tr.] 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     233 

It  is  to  be  hoped  that  an  extension  of  the  viewpoint  of  M.  H. 
Fischer,  together  with  that  of  A.  Oswald  will  explain  the  problem 
of  inflammation,  which  has  been  so  long  kept  at  a  dead  center. 

We  thus  see  that  the  science  of  colloids  throws  new  light  on  the 
most  difficult  portions  of  pathology,  and  that  even  therapy  may 
gaze  with  hope  upon  the  young  science. 

Salt  Distribution. 

Just  lis  the  water  content  of  a  normal  organ  is  relatively  constant 
and  may  undergo  reversible  changes  due  to  changes  in  the  condition 
of  the  organ  colloids,  this  is  also  true  in  the  case  of  the  salt  content. 

Unfortunately  the  basis  for  the  comprehensive  explanation  of 
these  questions  is  lacking  (bibliography,  see  Albu-Neuberg  *).  The 
values  that  have  been  obtained  cannot  be  used  for  comparison. 
Some  have  been  derived  from  healthy  and  some  (especially  the  de- 
terminations on  man)  from  sick  individuals;  moreover,  age  is  a  very 
important  factor.  In  the  first  place,  it  is  essential  to  determine 
accurately  the  limits  within  which  the  salt  content  in  the  separate 
organs  of  normal  individuals  may  vary.  We  know  that  muscle, 
liver,  blood  corpuscles  and  brain  are  rich  in  potassium  salts,  while 
in  the  blood  serum  and  spleen,  sodium  salts  predominate. 

In  man  the  muscles  contain  0.743  per  cent  of  NaCl,  the  lungs,  kid- 
neys and  skin  2.5  per  cent.  Sodium  chlorid  and  water  content  do  not 
run  parallel.  The  salt  content  varies  within  wide  Hmits  in  different 
species  of  animals.     For  instance,  there  is  present  in  the  ash  of 

Ox  blood 7 . 4  per  cent  K2O 

Calf  blood 11 . 2  per  cent  K2O 

Sheep  blood 7. 1  per  cent  K2O 

Pig  blood 20.4  per  cent  K2O 

Chicken  blood 18 . 4  per  cent  KoO. 

We  should  assume  from  our  colloid-chemical  knowledge,  that  a 
given  electrolyte  content  must  correspond  to  each  organ's  condition 
of  swelhng,  though  as  yet  this  assumption  offers  but  httle  towards 
elucidating  the  problem.  Pathological  retention  of  common  salt  in 
edema,  see  page  232,  invites  experimental  study  of  the  question. 

What  has  been  said  of  animals  is  also  true  of  plants.  But  here, 
too,  the  salt  content  varies  greatly  with  organ  and  sex,  and  we  have 
no  basis  for  its  true  significance.  For  instance,  it  is  merely  neces- 
sary to  mention  that  the  ash  of  wheat  flour  contains  0.76  per  cent 
Na20,  whereas  that  of  buckwheat  flour  has  5.87  per  cent  Na20. 

The  distribution  of  salts  occurs  similar  to  that  of  water;  if  they  are 
artifically  introduced  into  the  organism,  they  are  stored  up  and  again 
released.  If  a  dog  received  an  intravenous  injection  of  table  salt, 
28-77  per  cent  of  the  saline  retained  by  the  body  accummulates  in 


234  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

the  skin  (Padtberg).  In  salt  starvation  the  skin  acts  conversely, 
suffering  60-90  per  cent  of  the  chlorid  loss.  [The  ancient  name  for 
eczema  was  "salt  rheum";  see  also  "  Karell  Treatment,"  J.  G.  M. 
BuLLOWA,  Amer.  Medicine,  June,  1918.     Tr.] 

For  animals,  an  intravenous  injection  of  potassium  salts  acts  as 
a  poison  (especially  for  cardiac  muscle  and  peripheral  vessels); 
thus,  according  to  Held,  even  solutions  with  0.08  per  cent  KCl 
affect  frogs  and  rabbits.  By  way  of  the  intestinal  tract,  potassium 
salts  are  relatively  harmless.  Held  *  showed  that  when  thus  in- 
troduced, the  K  was  stored  in  the  tissues  and  only  slowly  given  up 
to  the  blood.  Here  we  observe  the  same  phenomena  as  with  water, 
of  which  an  excess  is  also  taken  up  by  the  tissues. 

For  the  maintenance  of  the  osmotic  pressure,  a  definite  concen- 
tration of  any  crystalloid  suffices.  Observations  of  the  most  differ- 
ent kind  teach  us  that  exactly  that  electrolyte  which  is  normally  found, 
is  necessary  for  function  and  development.  If  in  a  suspension  of 
blood  corpuscles  the  NaCl  is  replaced  by  sugar,  hemolysis  becomes 
more  difficult,  even  though  the  osmotic  pressure  is  identical.  Ex- 
periments on  excised  hearts  prove  that  the  action  of  the  heart  is  re- 
tained much  longer  if  it  is  perfused  with  a  fluid  containing  a  proper 
quantity  of  K,  Ca,  Mg,  PO4,  than  if  only  physiological  salt  solution 
is  used.  Na,  K,  Mg  and  Ca  salts  are  individually  poisonous  for 
plants,  but  mixed  in  the  proper  proportions  they  are  absolutely 
necessary.     In  Chapter  XXII,  there  are  further  examples. 

Evidently  the  condition  of  swelling  required  for  normal  function 
is  afforded  by  a  proper  balance  in  the  mixture  of  electrolytes. 

A.B.  Macallum  ^  investigated  microchemically  the  distribution  of  K, 
Fe,  Ca,  CI  and  PO4  in  many  animal  and  plant  cells,  and  from  his  investi- 
gation made  deductions  concerning  their  functional  significance,  to  which 
we  shall  again  return  (see  p.  292) .  Th.  Weevers  has  elaborated  them 
with  far  reaching  studies  of  the  distribution  of  potassium  in  plant  cells. 

[W.  BuRRiDGE,  Quarterly  Journal  of  Medicine,  10,  No.  39,  p.  172, 
aptly  remarks  that  analyses  of  the  blood  ash  give  little  information  con- 
cerning the  balance  of  its  salts  by  reason  of  the  fact  that  the  propor- 
tions of  them  which  are  in  "  sorption  "  or  in  solution  may  vary.     Tr.] 


1  A.  B.  Macallum  proceeds  in  part  from  the  fact  that  inorganic  salts  increase 
the  surface  tension  of  an  aqueous  solution,  and  as  a  result  the  surface  contains  a 
more  dilute  solution.  Conversely  the  author  concludes  that  the  surface  tension 
is  diminished  at  the  points  of  the  cells  which  are  approached  by  the  salts  in 
question.  We  cannot  agree  with  this  conclusion  at  present,  because  not  only 
mechanical  but  chemical  influences  may  determine  the  adsorption  of  salts.  For 
instance,  L.  Michaelis  and  P.  Rona  (as  the  author  has  mentioned)  demonstrated 
that  certain  kinds  of  sugar  have  no  influence  on  the  surface  tension  and  yet  may 
be  adsorbed. 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     235 

There  are,  moreover,  the  interesting  observations  of  P.  Rona  and 
D.  Takahashi,*  Hollinger*  and  E.  Frank*  concerning  the  dis- 
tribution of  sugar  between  blood  corpuscles  and  plasma. 

If  we  go  further  and  observe  the  distribution  of  the  other  ele- 
ments of  the  organism,  the  albiunins,  nucleins,  elastin,  the  lipoids, 
etc.,  we  approach  the  greatest  problems  of  anatomy  and  histology 
and  set  out  upon  a  boundless  and  uncharted  sea.  Possibly  we  shall 
learn  more  of  these  things  in  the  not  too  distant  future. 

The  Circulation  of  Material. 

Both  the  plant  and  the  animal  organism  are  surrounded  by  mem- 
branes or  pellicles,  which  separate  them  from  the  outer  world.  These 
membranes  are  more  or  less  permeable  to  water  and  crystalloids,  but 
normally,  they  are  impermeable  to  colloids.  Even  though  this  last 
fact  were  not  experimentally  demonstrated,  we  should  assume  it  a 
priori,  because  if  the  dissolved  colloids  could  leave  the  organism,  the 
loss  of  material  would  mean  death.  To  demonstrate  the  correct- 
ness of  our  conclusion,  we  need  but  mention  a  pathological  con- 
dition, albuminuria.  In  this  condition  the  kidney  becomes  permeable 
for  serum-albumin,  and  it  is  one  of  the  physician's  most  important 
duties  to  compensate  for  the  continuous  loss  of  substance  by  proper 
dieting,  so  that  no  impoverishment  of  the  tissues  as  regards  al- 
bumin occurs. 

Within  the  organism  also,  there  are  many  such  partitions;  they 
serve  to  organize  activity,  to  guide  the  food  along  certain  paths 
(arteries,  veins,  vascular  bundles  of  plants)  and  to  collect  secretions 
(urinary  bladder,  gall  bladder). 

The  substances  necessary  to  support  Hfe  must  accordingly  enter 
the  organism  as  gases  or  crystalloids.  In  the  case  of  plants,  CO2 
enters  through  the  leaves;  other  foodstuffs,  water  and  most  of  the 
inorganic  salts  (nitrates,  phosphates,  potassium  and  lime  salts, 
etc.),  enter  through  the  roots.  These  substances  are  at  the  outset 
very  diffusible  and  need  no  preparation.  It  is  otherwise  in  the  case 
of  animals,  which  require  outside  of  water  but  few  crystalloids 
(sugar,  salts)  and  are  chiefly  sustained  by  colloids  (vegetables  and 
meat).  In  order  to  enter  the  organism  at  all,  these  substances 
must  first  be  changed  to  a  crystalloidal  condition.  This  is  accom- 
plished by  enzymes;  the  diastatic  ferments  split  starches;  pepsin 
and  trypsin  split  protein;  and  herbivorse  have  ferments  which  are 
able  to  change  even  cellulose  into  a  crystalloidal  condition,  etc. 

In  like  manner  only  gases  or  crystalloids  can  leave  the  organism 
(expired  CO2,  urine,  perspiration). ^ 

1  Feces,  etc.,  do  not,  strictly  speaking,  leave  the  organism  any  more  than 
diatoms  which  have  been  surrounded  by  an  amoeba  and  then  cast  out  (they  are 
evacuated  from  a  tube  which  passes  through  the  animal). 


236  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  forces  which  accomphsh  the  entrance  of  food  into  the  organ- 
ism and  keep  up  the  circulation  of  ^natter  are  in  part  purely  mechani- 
cal, as  performed  by  the  lungs,  the  heart,  the  peristalsis  of  the 
intestines,  etc.  In  addition  to  these,  there  are  forces  which  ac- 
complish chiefly  the  metabolism  of  the  cells;  the  most  important  of 
these  are  diffusion,  osmotic  pressure,  swelling  and  shrinking.^ 

Circulation  of  Water. 

Until  a  few  years  ago  the  circulation  of  water  in  the  organism  was 
chiefly  attributed  to  osmosis.  The  vital  processes  constantly  produce 
from  the  colloids  osmotically  active  crystalloids,  which  both  retain  the 
water  formed  by  oxidation  and,  in  addition,  attract  water  into  the 
cells,  thus  maintaining  the  turgor  or  normal  tissue  tension.  This 
presupposes  that  an  almost  semipermeable  membrane  surrounds 
every  cell.  In  the  case  of  plants,  this  hypothesis  offers  certain 
difficulties,  and  in  the  case  of  animals,  it  is  impossible  to  maintain  it. 

We  shall  present  only  a  few  examples  which  show  that  osmotic 
conditions  alone  do  not  satisfactorily  explain  the  distribution  of 
water  in  animal  cells. 

Through  the  investigations  of  H.  J.  Hamburger,  H.  Koeppe  and 
E.  Overton,  it  is  known  that  in  the  presence  of  alterations  of 
osmotic  pressure,  blood  corpuscles  and  muscles  change  their  volume 
to  much  less  an  extent  than  would  be  expected  of  cells  with  fluid 
contents  and  a  semi-permeable  membrane.  Blood  corpuscles  contain 
about  60  per  cent  water.  In  his  experiment  with  osmosis.  Ham- 
burger showed  that  only  from  40  to  50  per  cent  of  their  volume 
could  consist  of  an  aqueous  solution,  so  that  from  10  to  20  per  cent 
of  the  water  arises  in  some  other  way.  According  to  Overton  the 
same  thing  holds  for  frog's  muscle. 

Water  is  also  retained  by  swelling.  Swelling  and  shrinking  are  the 
most  powerful  factors  governing  the  circulation  of  water  in  the 
organism.  They  may  even  act  against  osmotic  pressure;  nor  are 
we  forced  to  explain  their  activity  by  any  hypothetical  membranes. 
Changes  in  the  reaction  of  the  cells,  especially  the  constantly  recog- 
nizable acid  production  during  vital  processes,  give  rise  to  the  condi- 
tions necessary  for  swelling  or  the  circulation  of  water.  With  the 
removal  of  the  acids  shrinking  must  occur  again. 

M.  H.  Fischer  properly  cafls  attention  to  the  fact  that  a  semi- 
permeable membrane  permitting  the  entry  and  exit  of  water  from  the 

1  J.  Traube*!  regards  the  "surface  pressure"  as  the  force  which  causes  the 
movement  of  matter  in  the  organism.  Since  there  exists  a  certain  parallelism 
between  the  ability  of  many  substances  to  lower  surface  tension  and  their  capacity 
to  penetrate  the  cells,  Traube  disregards  the  osmotic  forces  and  Lipoid  solubility. 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL      237 

cell  is  a  monstrosity,  for  how  does  it  explain  the  entry  of  food  and  the 
exit  of  metabolic  products  (metabolites)  from  the  cell.  If  the  mem- 
brane is  permeable  for  these,  the  osmotically  active  crystalloids 
cannot  induce  transfer  of  water.  All  variations  in  volume  of  blood 
corpuscles,  spermatozoa,  plant  cells,  etc.,  produced  by  electrolytes 
and  attributed  to  osmotic  pressure  up  to  now,  are  just  as  well  ex- 
plained by  swelling  and  shrinldng.  Gels  swell  up  in  water,  acids  and 
alkalies;  salts  on  the  other  hand,  hinder  sweUing  and  cause  shrinkage. 
Moreover,  new,  purely  physico-chemical  observations  likewise  warn 
us  to  employ  great  caution  in  our  consideration  of  osmotic  processes 
in  the  organism.  Much  more  substance  may  be  dissolved  in  the  in- 
terior of  the  cell  than  in  its  surrounding  fluid  without  the  osmotic 
pressure  making  this  evident.  We  saw  on  pages  46  and  47  that 
with  decrease  in  the  surrounding  osmotic  pressure,  the  osmotic 
pressure  in  a  cell  with  a  permeable  membrane  containing  colloid, 
falls,  although  if  reckoned  according  to  the  salt  content,  the  osmotic 
pressure  should  have  increased.  These  findings  of  W.  Biltz  and  A. 
VON  Vegesack*  necessitate  a  revision  of  all  former  conclusions 
derived  from  the  observation  of  osmosis  in  cells. 

Circulation  of  Water  in  Animals. 

A  movement  of  water  results  when  conditions  arise  which  change 
the  relative  swelling  of  the  organs.  When  subjected  to  high  tem- 
perature or  after  violent  exercise,  etc.,  the  skin  loses  water,  the 
blood  loses  water  through  the  lungs,  which  causes  a  flow  of  water 
from  the  other  organs.  Conversely,  an  excess  of  water  from  the  in- 
testines, or  in  the  case  of  frogs  and  certain  other  animals  from  the 
skin,  is  transferred  to  other  organs  and  re-excreted  by  the  kidneys. 
Other  circumstances  may  arise,  however,  which  determine  the  cir- 
culation of  water:  concentration  of  acid  in  a  tissue  increases  its 
swelling  capacity,  attracting  water,  e.g.,  in  venous  blood  or  an  edema, 
whereas  simultaneous  salt  formation  leads  to  a  shrinking  or  loss  of 
water.^  In  circumstances  in  which  osmotic  pressure  may  become 
active,  as  when  a  membrane  is  interposed,  the  change  of  a  colloidal 
substance  into  a  crystalloid  under  the  influence  of  enzymes  may  effect 
a  transfer  of  water;  the  water  flows  to  the  place  where  the  osmotic 
pressure  is  higher.  We  shall  return  to  the  details  of  this  question 
when  we  consider  the  individual  organs. 

'  From  this  it  results  that  the  presence  of  colloids  regulates  the  movement  of 
water  in  an  entirely  different  and  at  times  in  a  direction  opposite  to  that  of  the 
osmotic  pressure:  Acid  +  salt,  as  a  result  of  the  higher  osmotic  pressure,  should 
increase  the  amount  of  water  attracted;  in  the  case  of  colloid  structures,  how- 
ever, they  decrease  it,  since  salts  aboUsh  to  a  greater  or  less  extent  the  swelling 
action  of  acids. 


238  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  Movement  of  Water  in  Plants. 

The  evaporation  of  water  in  plants  proceeds  more  rapidly  than 
in  animals.  The  enormous  development  of  surface  in  the  shape  of 
leaves  and  needles  underlies  a  great  transpiration  which  requires 
replacement,  so  that  a  stream  of  water  moves  upward  through  the 
roots  and  vascular  bundles  to  the  leaves.  On  bright  summer  days 
(see  W.  Pfeffer  *2  loc.  cit.  I,  p.  233)  1  to  10  gm.  water  are  evaporated 
from  1  cm.2  of  leaf  surface.  On  very  hot  days  the  loss  by  transpiration 
from  big  trees  exceeds  400  kilos;  on  rainy  days,  however,  it  may  be 
reduced  to  a  few  kilos.  To  explain  the  upward  movement  of  the 
water,  the  most  varied  theories  have  been  advanced,  and  usually 
abandoned.  Explanation  by  means  of  osmotic  pressure  has  proved 
thoroughly  unsatisfactory,  and  mere  capillary  imbibition  is  of  no 
greater  use,  We  may  well  understand  that  the  colloids  of  leaves 
suffer  a  loss  of  water  by  evaporation,  and  that,  in  swelling,  they  are 
able  to  lift  a  great  column  of  water  from  the  ground  to  the  tree  top. 
Experiments  of  E.  Strassburger  showed  that  in  poisoned  trees, 
water  may  rise  to  a  height  of  22  meters,  so  that  pure  capillary  forces 
do  not  suffice  for  the  explanation  of  the  phenomena.  More  recent  ex- 
periments (P.  A.  RosHARDT,*  E.  Reinders*)  show  that  in  the  living 
plant,  living  elements  assist  in  pumping  up  the  water.  Since  no  pre- 
vious explanation  of  this  has  been  given,  I  believe  that  I  am  justified 
in  formulating  the  following  hypothesis.  In  my  opinion,  the  living 
cells  of  plants  assist  in  the  elevation  of  the  sap  by  their  respiration. 
With  respiration,  not  only  does  CO2  develop,  but  also  great  quantities 
of  organic  acids.  Both  cause  a  swelling  or  attraction  of  water,  which 
is  liberated  to  the  extent  that  CO2  disappears,  and  the  other  acids  are 
removed  in  any  one  of  the  many  possible  ways.  This  would  fit  in 
with  the  fact  that  the  breathing  in  fully  developed  leaves  and 
branches,  in  which  the  need  for  water  is  also  diminished,  is  less  than 
in  the  developing  shoots.  The  dead  leaf,  whose  breathing  has  ceased, 
withers. 

Circulation  of  Crystalloids. 

The  circulation  of  crystalloids  is  also  largely  governed  by  the  factors 
of  diffusion  and  osmotic  pressure,  with  certain  limitations  due  to 
the  colloid  media.  Although  between  two  aqueous  solutions,  sep- 
arated by  an  easily  permeable  membrane,  unrestricted  mixing  occurs 
as  a  result  of  diffusion,  this  does  not  hold  for  a  jelly-like  medium 
(see  H.  Bechhold  and  J.  Ziegler*^).  In  order  to  bring  about  a 
mixture  in  such  a  case  an  excess  of  osmotic  pressure  is  required 
(see  p.  57).     It  even  seems  that  with  equal  osmotic  pressure,  acid 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     239 

and  alkaline  reacting  substances  may  lie  side  by  side  in  colloidal 
(amphoteric)  media  for  a  long  time  without  neutralizing  each  other 
(R.  E.  Liesegang).  In  the  case  of  phagocytes,  that  is,  in  Hving  cells, 
the  existence  of  acid  areas  in  alkaline  protoplasm  has  been  shown  by 
staining  with  neutral  red  (E.  Metschnikoff).  We  thus  see  that  in 
different  portions  of  the  organism,  the  most  various  crystalloids  are 
present,  and  may  functionate  specifically  without  being  accompanied 
by  any  exchange  or  mixture;  this  only  occurs  when  a  crystalloid  sub- 
stance accumulates  and  becomes  osmotically  active.  Swelling  and 
shrinking  may  also  be  of  importance  for  the  circulation  of  crystal- 
loids, since  dissolved  substances  are  soaked  up  with  the  water  of 
swelling  or  are  expressed  during  shrinking. 

If  these  crystalloids  are  at  the  same  time  electrolytes,  they  may 
increase  or  diminish  the  swelling  according  to  their  nature  (acid  or 
salt);  and  in  this  way,  either  aid  or  impede  the  entrance  of  crystal- 
loids. 

Circulation  of  Colloids. 

Compared  with  crystalloids,  the  osmotic  pressures  in  the  case  of 
colloids  are  extremely  small.  To  be  sure,  we  know  (see  p.  55)  that 
proteins  may  diffuse  through  gels,  so  that  they  also  are  independ- 
ently motile.  Of  great  significance  is  the  discovery  of  H.  Iscovesco 
to  the  effect  that  colloid  diffusion  is  dependent  on  the  electric  charge. 
In  general,  however,  the  colloids,  as  opposed  to  the  crystalloids, 
furnish  the  stable  element  of  the  organism. 

The  Influence  of  Membranes  Upon  the  Interchange  of  Substances. 

The  physico-chemical  conditions  for  the  interchange  of  substances 
through  cell  membranes  was  for  a  long  time  completely  ruled  by  the 
theory  of  Overton,  which  is  somewhat  as  follows:  Protoplasm  is 
surrounded  by  a  fatty  lipoid  membrane;  an  exchange  of  substances 
can  only  occur  if  the  given  substance  is  soluble  in  such  a  membrane. 
Overton's  theory  has  not  proven  universally  applicable;  it  is  ever 
becoming  better  recognized  that  the  problem  will  probably  be  solved 
when  we  cease  to  look  entirely  to  the  osmotic  conditions  and  mem- 
branes for  the  factors  governing  the  interchange  of  substance.  The 
fact  that  both  cell  content  and  cell  membrane  consist  of  colloids 
capable  of  swelling  must  be  taken  into  consideration. 

The  earliest  fundamental  investigations  of  the  physical  inter- 
change of  matter  in  individual  cells  were  made  on  plant  cells.  I 
refer  particularly  to  the  investigations  of  W.  Pfeffer  and  H.  de 
Vries.     In  plants,  especially,  we  find  that  the  cell  content  is  very  fre- 


240  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

quently  surrounded  by  a  visible  and  solid  membrane  which  is  usually 
regarded  as  semipermeable. 

The  basis  of  this  view  is:  if  such  a  cell  is  placed  in  hypertonic 
salt  solution,  the  protoplasm  retracts  from  the  cell  wall  and  water  is 
lost.  This  phenomena  is  called  plasmolysis.  If  the  cell  is  im- 
mersed in  pure  water,  the  protoplasm  swells  up  again.  The  phe- 
nomenon was  formerly  explained  by  saying  that  the  membrane  was 
impermeable  for  salts.^  In  a  hypertonic  salt  solution,  water  may  in- 
deed leave  the  cell  but  salt  cannot  enter;  in  pure  water  the  process 
is  reversed. 

Nowadays  discussion  is  focussed  on  the  nature  of  the  plasma 
pelHcle  and  two  main  tendencies  may  be  recognized. 

Among  the  adherents  to  the  lipoid  theory  in  addition  to  E.  Oveeton, 
is  Vernon,  who  considers  it  probable  that  the  hpoid  membrane 
penetrates  the  interior  of  the  cell.  J.  Loeb  and  R.  Beutner  in  view 
of  their  investigations  of  bio-electric  phenomena  may  be  regarded  as 
adherents  of  the  Hpoid  theory.  Those  investigators  (J.  Traube  and 
F.  Czapek)  who  regard  changes  in  surface  tension  as  the  means  of 
penetrating  surfaces  may  be  regarded  as  adherents  of  a  modified 
lipoid  membrane  theory.  They  arrive  at  this  conclusion  because 
their  experiments  have  been  chiefly  concerned  with  the  action  of 
lipoid  soluble  substances  on  the  cell. 

According  to  F.  Czapek  all  substances  whose  surface  tension  is 
less  than  0.68  (water/air  =  1)  are  toxic  for  the  higher  plant  cells  and 
Czapek's  pupil  KiscH  determined  0.5  to  be  the  hmit  of  toxic  surface 
tension  for  yeast  cells  and  fungi.  Since  lecithin  and  Cholesterin,  that 
is,  the  lipoids  and  their  emulsions,  have  a  surface  tension  of  0.5, 
F.  Czapek  agrees  with  Nathanson  and  regards  the  cell  membrane 
as  a  concentrated  fat-emulsion  which  is  permeable  for  either  fat  or  for 
water  soluble  substance  depending  on  the  conditions  of  surface  ten- 
sion. Similar  views  (loose  union  of  albumin  and  lipoid)  are  enter- 
tained by  W.  W.  Lepeschkin  with  the  difference  that  he  regards  the 
entire  protoplasm  as  such  an  emulsion  possessing  properties  in  the 
center  similar  to  those  on  the  surfaces. 

The  "emulsion  theory"  obtained  very  definite  support  from  the  fol- 
lowing observation  of  Clowes  (see  p.  38).  He  prepared  an  oil-water 
emulsion  by  shaking  equal  quantities  of  water  and  olive  oil  and  suffi- 
cient n/10  NaOH  that  the  outer  phase  (the  water)  was  just  alkahne 
to  Phenolphthalein,  If  he  now  added  a  small  excess  of  CaCl2  solution 
the  emulsion  changed  into  a  water-oil  emulsion;  in  other  words, 
water  became  the  dispersed  phase  in  a  continuous  layer  of  oil.    We 

1  The  visible  cell  membrane  is  quite  permeable  for  most  crystalloids,  serving 
only  to  a  certain  extent  as  a  support  for  the  protoplasm. 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     241 

observe  that  by  this  chemical  attack  the  layer  which  had  been  per- 
meable for  hydrophile  substance  became  impermeable  for  them  and 
was  made  permeable  for  substances  soluble  in  fat.  However,  we  know 
from  the  investigations  of  J.  Loeb  and  W.  J.  V.  Osterhout  (see  p. 
378,  et  seq.)  that  small  amounts  of  divalent  cations  detoxicate  neutral 
salts  by  inhibiting,  according  to  the  view  of  J.  Loeb,  the  free  ex- 
change of  ions  through  the  plasma  pellicle.  Clowes  extended  his 
observations  to  other  polyvalent  cations  and  the  quantitative  rela- 
tions are  in  excellent  agreement. 

The  other  tendency  is  to  assume  a  pure  albuminous  membrane; 
W.  J.  V.  Osterhout  assumes  this,  as  the  result  of  the  following 
remarkable  observation:  he  placed  spyrogyra  cells  in  common  salt 
solution  of  such  concentration  that  no  plasmolysis^  occurred;  when 
he  added  very  dilute  calcium  chlorid  solution  so  as  to  depress  the 
osmotic  pressure,  plasmolysis  occurred.  The  plasma  pellicle  must 
have  been  permeable  for  NaCl  and  its  passage  is  only  impeded  by 
the  CaCl2.  In  contrast  to  Overton's  view,  Osterhout  regards  the 
plasma  pellicle  as  permeable  for  most  ions  of  the  light  metals  and 
consequently  it  must  be  albuminous. 

The  action  of  the  Ca-ion  possibly  depends  on  its  antagonistic 
action  (see  p.  69)  though  it  may  be  due  to  a  variety  of  tannage  of 
the  plasma  pellicle.  With  the  death  of  the  cell,  the  pellicle  becomes 
generally  permeable. 

RuHLAND  also,  discards  the  lipoid  theory.  He  considers  only  the 
thickness  of  the  membrane  to  be  responsible  for  permeability  or 
impermeability;  the  membrane  acts  like  an  ultrafilter  in  the  sense  of 
Bechhold.  He  studied  a  large  number  of  dyes,  enzymes,  alkaloids 
and  other  substances  which  occur  in  plants  and  found  that  their 
ability  to  penetrate  the  plasma  cells  was  in  proportion  to  their  ability 
to  spread  out  in  thick  jellies;  in  other  words,  it  depended  on  their 
particle  size  (see  p.  56). 

In  view  of  the  known  facts  we  must  admit  that  at  present  we  can 
arrive  at  no  conclusion  concerning  the  nature  and  structure  of  the 
plasma  pellicle.  Of  one  thing  we  can  be  certain,  that  Overton's 
original  theory  of  a  continuous  lipoid  membrane  must  be  abandoned. 
I  am  of  the  opinion,  however,  that  it  is  possible  to  conciliate  the 
theories  which  have  been  elucidated  here  and  which  seem  to  be 
mutually  exclusive. 

In  the  first  place,  the  assumption  of  a  pellicle  of  emulsified  fat  does 

^  Osterhout  distinguished  between  true  and  false  plasmolysis.  The  latter 
may  occur  in  dilute  solutions  even  in  pure  water  most  usually  in  marine  plants. 
It  is  probably  due  to  the  coagulation  of  the  protoplasm  from  the  penetration  of 
the  water. 


242  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

not  exclude  Ruhland's  ultrafilter  theory.  If  we  have  an  emulsion 
in  which  the  lipoid  is  the  dispersed  phase  we  have  an  ultra  filter  which 
is  permeable  for  water  soluble  substances  and  impermeable  for  lipoid 
soluble  substances.  The  size  of  the  pores  of  the  ultra  filter  depends 
on  the  relation  of  the  lipoid  to  the  aqueous  phase.  If  the  amount  of 
lipoid  is  small,  the  pores  of  the  ultra  filter  are  large  and  vice  versa. 
When  the  lipoid  content  is  large  we  have  a  narrow  pored  ultra  filter 
which  absolutely  satisfies  the  conditions  found  by  Ruhland  in  his 
dye  investigations.  Further,  we  have  seen  from  Clowes'  experiment, 
that  an  oil/water  emulsion  is  easily  changed  to  a  water/oil  emulsion, 
and  in  that  case  the  layer  is  open  for  fat  soluble  substances  and  closed 
for  water  soluble  substances  In  my  opinion  such  a  layer  satisfies 
all  the  conditions  demanded  by  the  various  investigators. 

I  wish,  however,  to  emphasize  that  there  is  no  justification  for  too 
wide  a  generalization,  for  different  cells  behave  very  differently. 
Observations  on  plant  cells  cannot  be  applied  without  modification 
to  animal  cells;  a  cell  in  a  plant  root  cannot  be  compared  to  nerve 
cells  which  are  surrounded  by  a  dense  isolating  layer  of  fat.  It  seems 
possible  to  conclude  from  R.  Höber's  and  Ruhland's  experiments 
on  the  penetration  of  dyes  into  cells  that  the  animal  cells  which  they 
studied  contain  larger  pores  than  the  plant  cells. 

Let  us  consider  the  simplest  instance,  one  in  which  the  cell  proto- 
plasm is  a  colloid  capable  of  swelling,  with  surfaces  limited  by  a 
pellicle  which  can  also  swell  and  offering  certain  exterior  boundaries. 
Any  injury  to  this  pellicle  will  be  repaired  of  its  own  accord  somewhat 
like  rubber.  We  can  thus  (pp.  284-285)  readily  understand  how 
amoeboe  or  phagocytes  send  out  protoplasmal  prolongations,  envelop 
foreign  bodies  or  bacteria  and  incorporate  them  without  their  margin 
being  broken.  As  a  matter  of  fact,  it  must  immediately  repair  itself 
just  as  does  an  oily  film  on  water  broken  by  a  stone.  Let  us  see  how 
this  view  agrees  with  former  theories,  and  to  what  extent  this  view  is 
an  improvement  upon  them. 

To  begin  with,  it  must  be  noted  that  Overton  assumes  that  a  sub- 
stance is  taken  up  by  the  plasma  pellicle  in  accordance  with  its 
coefficient  of  solubility,  in  agreement  with  the  laws  of  solutions 
(Henry's  distribution).  This  may  be  the  fact  in  many  cases,  only  we 
must  recall  that  adsorption  fulfills  similar  conditions  for  the  passage 
of  a  substance  through  the  plasma  pellicle  into  the  interior  of  the 
cell.  The  only  condition  which  need  be  assumed  in  order  that  a 
substance  may  enter  the  interior  of  a  cell  from  outside,  is  that  there 
shall  be  a  reversible  absorption  by  the  plasma  film.  What  curve  of 
distribution  this  follows,  is  immaterial  for  the  present.  That,  as  a 
matter  of  fact,  in  numerous  cases  an  adsorption  certainly  does  exist, 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     243 

but  not  a  distribution  according  to  Henry's  law,  has  been  determined 
by  H.  Bechhold  in  the  action  of  disinfectants  (see  p.  399),  and 
Straub-Freundlich  on  the  distribution  of  veratrin  between  heart 
muscle  and  pericardial  blood.  G.  Loewe  has  shown  by  simple 
physico-chemical  experiments  that  lipoids  adsorb  dyes,  narcotics, 
nicotin  and  tetanus  toxin.  The  substance  interchange  in  animal  cells 
has  been  studied  most  thoroughly  in  the  case  of  red  blood  corpuscles. 
In  my  opinion  (see  p.  304)  the  latter  have  a  very  peculiar  struc- 
ture, conditioned  by  their  special  function;  their  lipoid  pellicle 
is  quite  strong.  In  spite  of  this,  we  shall  find  phenomena  in 
the  case  of  the  erythrocytes  which  cannot  be  brought  into  accord 
with  the  idea  of  a  salt  solution  surrounded  by  a  semipermeable 
membrane. 

The  theory  of  osmotic  pressure  demands  that  various  isotonic  salt 
solutions  shall  have  equal  influence  upon  the  volume  of  the  blood 
corpuscles.  S.  G.  Hedin*^  showed,  however,  that  this  is  not  the 
case,  for  instance,  in  isotonic  solutions  of  NaCl  and  KNO3;  in  the  case 
of  lower  concentrations,  the  volume  is  smaller;  in  the  case  of  higher 
concentrations  it  is  larger  than  with  the  corresponding  NaCl  solu- 
tion; we  must  recall  that  the  NO3  ion  favors  swelling  or  the  de- 
flocculation  of  colloids  and  lecithin;  if  the  outer  pressure  is  low, 
crystalloids  leave  the  blood  corpuscles,  and  the  osmotic  pressure,  and 
consequently  the  volume  of  the  corpuscles,  will  be  less  than  with  the 
corresponding  NaCl  solution.  The  reverse  occurs  if  the  outer  solu- 
tion is  hypertonic. 

We  find  in  the  literature,  repeated  references  to  the  permeability 
of  the  cell  membrane,  especially  of  plants,  for  'potassium  nitrate.  B. 
VAN  Rysselberghe*  has  demonstrated  the  entrance  of  diphenylamin 
into  tradescantia  cells.  If  fungi,  such  as  aspergillus  niger  or  penicil- 
lium  glaucum,  are  grown  upon  a  concentrated  solution  of  saltpeter, 
they  will  take  up  so  much  of  the  electrolyte  that  in  the  end  they  will 
have  an  osmotic  pressure  of  200  atmospheres.  Such  cultures  actually 
explode  when  placed  in  pure  water. 

The  ability  to  take  up  such  substances  as  favor  swelling  is  much 
greater  in  the  case  of  young  cells  with  membranes  that  can  swell 
than  in  the  case  of  old  inelastic  cells.  On  this  account,  an  older 
aspergillus  cell  may  plasmolyze  with  a  20  per  cent  NaNOs  solu- 
tion which  possesses  an  osmotic  pressure  of  only  102  atmospheres. 
In  this  difference  between  old  and  young  cell  membranes,  may  lie  a 
partial  explanation  why  bacteria  and  fungus  cultures,  namely  organ- 
isms which  multiply  rapidly,  readily  adapt  themselves  to  changed 
conditions.     Young  and  old  cells  differ  in  their  turgidity. 

R.  HöBER*^  prepared  suspensions  of  blood  corpuscles  in  dilute  iso- 


244  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

tonic  solutions  of  various  alkali  salts,  and  observed  the  order  in  which 
they    favored    hemolysis.      He    established    the    following    series: 

S04<  CI  <  Br,     NO3  <  I,     and    Li,  Na  <  Cs  <  Rb  <  K. 

The  anion  series  corresponds  fairly  well  with  the  action  of  the  anions 
upon  lecithin,  so  that  we  may  safely  assume  that  alkali  salts  may 
bring  about  an  increase  or  a  diminution  in  the  porosity  of  the  plasma 
pellicle.  Other  examples  of  this  action  on  the  part  of  neutral  salts 
are  given  in  Chapter  XXII  (Salts). 

Substances  which  favor  swelling  in  a  diluted  condition  (p.  68) 
may  prevent  it  when  they  are  more  concentrated.  It  must  also  be 
borne  in  mind,  that  besides  hydrophile  lecithin  and  albumin,  hydro- 
phobe Cholesterin  must  also  exist  in  the  lipoid  membrane.  This  lat- 
ter is,  however,  precipitated  by  electrolytes,  which  cause  the  former 
to  swell.  Thus  there  exists  in  the  cell  membrane  a  self -regulating 
system,  something  like  a  compensation  pendulum;  when  the  tem- 
perature rises,  the  center  of  gravity  of  the  pendulum  falls,  but  by  a 
combination  of  metalhc  rods  the  center  of  gravity  is  raised  and 
the  fall  is  compensated.  This  compensatory  action  of  hydrophile 
and  hydrophobe  colloids  appears  to  be  an  essential  factor  in  the 
automatic  regulation  of  cell  metabolism. 

The  following  considerations  afford  an  explanation  for  some  par- 
ticular kinds  of  cells.  R.  Höber*^  properly  calls  attention  to  the 
fact  that,  ''the  plasma  film  is  really  impermeable  for  everything 
the  cell  needs  or  produces."  It  is  impermeable  for  amino  acids, 
for  the  various  kinds  of  sugar  and  soluble  carbohydrates  which  are 
formed  in  the  interior  of  a  cell  from  the  undissolved  carbohydrate 
reserves  and  for  inorganic  salts  and  salts  of  organic  acids.  The  in- 
terchange of  these  substances,  which  naturally  rnust  occur,  is  on  this 
account,  somewhat  of  a  riddle.  In  my  opinion,  these  phenomena 
are  less  mysterious  if  we  recall  that  with  equal  osmotic  pressure 
without  and  within,  even  the  thinnest  membranes  interfere  with 
diffusion,  as  has  been  shown  by  H.  Bechhold  and  J.  Ziegler  (see 
p.  57).  The  most  recent  investigations  of  H.  J.  Hamburger  on 
blood  corpuscles  indicate  that  their  transition  membrane  is  not  as 
impermeable  as  was  formerly  believed.  It  will  be  understood  thus 
how  the  cells  are  sharply  cut  off  in  case  of  isotonicity,  while  if  there 
be  hypertonicity,  some  substance  may  penetrate  through  the  mem- 
brane. This  seems  to  me  to  be  the  meaning  of  the  following  ex- 
periment of  J.  Bang*  :  if  red  blood  corpuscles  are  placed  in  an  8  per 
cent  cane  sugar  solution  and  the  solution  is  immediately  diluted, 
hemolysis  occurs  when  the  cane  sugar  concentration  is  5.4  per  cent. 
If,  however,  the  red  blood  corpuscles  remain  in  the  cane  sugar  solu- 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     245 

tion  for  several  hours  and  the  dilution  is  then  undertaken,  the 
hemolysis  will  occur  only  when  the  cane  sugar  concentration  reaches 
2  per  cent.  Sufficient  time  has  thus  been  given  for  salts  to  leave 
the  blood  corpuscles  and  enter  the  cane  sugar  as  was  shown  by  A. 

GiJRBER. 

Experiments  of  Jaques  Loeb*'*  upon  the  parthenogenesis  of  sea 
urchin's  eggs  are  in  accord  with  this.  If  the  eggs  are  placed  for  a 
short  time  in  hypertonic  salt  solution  and  then  returned  to  sea 
water  (which  corresponds  with  their  normal  osmotic  pressure)  seg- 
mentation takes  place.  J.  Loeb**  found  that  a  cane  sugar  solution 
acts  like  a  hypertonic  salt  solution  even  if,  as  regards  concentration, 
it  be  isotonic  with  the  eggs.  J.  Loeb  explains  the  action  by  saying 
that  the  egg  pellicle  is  permeable  for  sugar  and  salts,  and  that  the 
salts  diffuse  out  more  rapidly  than  the  cane  sugar  diffuses  in,  so  that 
the  outer  fluid  becomes  hypertonic.  We  must,  moreover,  recall  that 
substances  exist  which  to  a  certain  extent  close  the  pathways  auto- 
matically, as  for  instance  the  SO4  ion,  whereas  others,  especially  urea, 
open  a  passage,  not  only  for  themselves  but  for  other  substances 
(see  p.  55).  The  permeability  of  red  blood  corpuscles  and  muscles 
for  urea  is  then  no  longer  surprising,  any  more  than  the  changes  in 
permeabihty  (observed  by  M.  Fluri*  and  R.  Meurer*)  in  the 
plasma  pellicle  of  plants  under  the  influence  of  certain  salts. 

This  may  be  accomplished  not  only  by  chemical  agencies,  but 
purely  physical  factors  may  have  an  influence.  It  might  be  ex- 
pected a  priori  from  change  in  temperature;  the  influence  of  light  is 
surprising,  as  experiments  of  W.  W.  Lepeschkin  and  by  A.  Tröndle 
have  shown.  The  latter's  experiments  indicate  that  plant  cells 
(foliage)  are  more  permeable,  not  only  for  NaCl  but  even  for  glucose, 
in  a  bright  light  than  in  the  dark. 

One  of  the  most  remarkable  and  still  unexplained  phenomena  is 
that  when  death  occurs,  the  permeability  of  the  cell  membrane 
changes  into  that  of  an  ordinary  membrane  which  retains  only  col- 
loids. 

Assimilation  and  Dissimilation. 

After  a  crystalloid  foodstuff  has  entered  the  organism,  it  is  the 
organism's  most  important  task  to  retain  it  for  use;  this  is  ac- 
complished by  changing  it  into  a  colloid,  inasmuch  as  complicated 
combinations  are  formed  from  more  or  less  simply  constructed  crys- 
talloids. From  the  CO2,  which  enters  the  leaf,  starch  is  formed 
under  the  influence  of  chlorophyl  granules  and  daylight;  and  from 
nitrates  which  have  entered  through  the  roots,  with  the  assistance 
of  carbohydrates,  proteins  develop.     In  the  animal  organism,  the 


246  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

readily  diffusible  peptones  which  have  entered  through  the  intestine 
are  again  changed  even  in  the  intestinal  wall  into  colloidal  albumin; 
sugar  is  retained  in  the  liver  as  an  animal  starch  or  glycogen  and  so 
on  for  other  examples. 

This  change  into  the  colloidal  condition  is  usually  associated  with 
a  metamorphosis  into  a  substance  native  to  the  body  from  which 
the  cells  and  tissues  of  the  organism  are  built  up.  It  may  be  con- 
cluded from  the  investigations  of  Alexis  Carrel  and  Burrows* 
that  the  circulating  nutritive  fluid  already  contains  all  the  elements 
required  for  the  most  varied  organs.  These  investigators  suspended 
pieces  of  tissue  from  freshly  killed  mammals  in  drops  of  plasma  from 
the  same  kind  of  animal.  The  tissues  continued  to  grow,  cartilage 
produced  cartilage,  a  spleen  produced  cells  which  resembled  spleen 
pulp,  and  pieces  of  kidney  grew  tubes  of  cells  which  resembled  the 
kidney  tubules. 

The  future  will  teach  us  whether  we  must  regard  this  phenomenon 
as  a  kind  of  crystallization.  Perhaps  it  will  be  possible  for  future 
colloid  investigators  to  express  the  problem  of  cell  nutrition  in  terms 
of  the  brilliant  side-chain  theory  of  Ehrlich.  To  what  extent  the 
fixation  of  colloid  foodstuffs  by  the  cell  is  a  matter  of  chemical 
forces  or  simple  mechanical  adsorption  is  an  open  question,  which 
so  far  must  be  decided  differently  in  each  individual  case. 

Hitherto  it  has  been  only  possible  in  the  case  of  fats,  to  follow 
visibly  their  course  from  the  moment  of  resorption  to  their  fixation 
in  body  tissues.  In  the  intestines^  with  the  assistance  of  the 
alkaline  reaction  of  the  intestinal  fluids,  the  intestinal  and  pan- 
creatic juices  and  the  bile  form  a  very  fine  emulsion  of  the  fats. 
Simultaneously,  there  occurs  a  splitting  into  fatty  acids  and  glycerin 
under  the  influence  of  ferments,  lipases.  It  is  not  yet  established 
whether  the  splitting  of  the  fats  is  complete,  which  would  mean  that 
the  intestine  could  only  absorb  dissolved  fatty  acid  salts  of  alkalis 
(soaps)  and  glycerin,  or  whether  some  of  the  fat  remains  unchanged 
and  is  absorbed  as  such.  If  this  latter  statement  is  actually  true,  we 
should  have  to  assume  that,  under  the  influence  of  the  surrounding 
soap  solution  and  possibly  other  factors,  the  surface  tension  of  the 
fat  droplets  is  reduced  to  a  minimum  so  that  they  may  easily  change 
its  shape  or  enlarge  their  surface  and  pass  the  very  minute  openings 
in  the  intestinal  epithelium.  From  page  16,  we  know  that  to  produce 
a  change  of  form  by  pressure  alone,  there  would  be  required  forces 
(many  atmospheres)  such  as  never  occur  in  the  organism.  We  have 
in  this  case  conditions  similar  to  those  in  the  case  of  leucocytes  (see 

1  It  is  hardly  possible  to  attribute  great  significance  to  saponification  in  the 
stomach. 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     247 

pp.  283  to  286),  which,  in  spite  of  their  considerable  diameter,  change 
their  shape  so  that  they  pass  through  the  finest  vessel  walls.  If  to  the 
leucocytes  is  to  be  attributed  a  share  in  the  resorption  of  fat,  they 
must  journey  into  the  intestines  and  return  laden  with  fat.  It  has 
not  been  possible  as  yet  to  decide  microchemically  whether  fat  passes 
the  intestinal  epithehum  unchanged.  [In  the  presence  of  protective 
colloids,  colloidal  gold  will  pass  through  Pukall  filters  which  other- 
wise hold  them  back.  Zsigmondy-Alexander,  Colloids  and  the 
Ultramicroscope,  p.  153,  et  seq.     Tr.j 

One  fact,  at  least  to  me,  seems  very  much  to  favor  the  idea  that 
fat  may  be  resorbed  unchanged  from  the  intestines,  namely,  linseed 
oil,  sesame  oil,  cottonseed  oil,  etc.,  may  occur  unchanged  in  the 
milk,  and  foreign  fats  (rapeseed  oil)  may  be  deposited  in  the  body. 
Absorption  occurs  almost  exclusively  in  the  small  intestines  (Naka- 
shima).  Within  the  intestinal  wall,  neutral  fat  may  be  synthesized 
from  the  absorbed  fatty  acid  alkali  and  glycerin,  so  that  neutral 
fat  is  carried  to  the  body  in  very  fine  emulsion  through  the  chyle 
ducts,  and  in  fact  fat  may  enter  the  blood  stream  directly. 

The  milky  turbid  lymph  collects  in  the  thoracic  duct  and  empties 
into  the  subclavian  vein.  It  is  especially  easy  after  the  ingestion  of 
fat  to  recognize  ultramicroscopically,  in  the  blood,  numerous  gran- 
ules (hemoconia),  which  may  be  considered  fat  droplets  (A.  Neu- 
mann,* K.  Reicher*). 

It  is  possible,  therefore,  to  follow  visually  the  path  of  fat  by  means 
of  dark  field  illumination.  S.  Bondi  and  A.  Neumann*  experimented 
as  follows :  at  times,  they  caused  hemoconia  to  appear  in  the  blood 
by  a  liberal  fat  diet,  and  at  others  they  injected  a  very  fine  emulsion 
of  fat  into  the  veins.  Large  fat  droplets  suspended  in  blood  are  evi- 
dently unable  to  change  their  form,  and  as  a  result  cause  emboli  in 
the  lungs,  which  may  prove  fatal.  These  investigators  experimented 
thus  with  emulsions  of  lanolin,  Cholesterin,  lecithin,  butter  and 
olive  oil,  whose  particles  were  only  recognizable  in  the  dark  field. 
They  dissolved  the  fat  they  were  using  in  alcohol  and  poured  the 
alcoholic  solution  slowly  into  water  with  constant  stirring.  The 
alcohol  was  removed  from  the  filtrate  of  this  emulsion  by  gently 
warming  it  on  a  water  bath. 

S.  Bondi  and  A.  Neumann  then  established  that  the  fat  droplets 
were  not  dissolved  by  lipolytic  ferments  during  their  sojourn  in  the 
blood.^  They  are  emulsified  by  the  venous  blood  in  the  right  heart, 
and  after  they  have  passed  the  capillaries  of  the  lesser  circulation, 
they  enter  those  of  the  greater  circulation.     Their  goal,  like  that  of 

^  In  my  opinion  such  emulsions  of  uniform  particle  size  could  serve  in  measur- 
ing the  exact  dimensions  of  the  smallest  capillaries  under  normal  conditions. 


248  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

other  suspensions  (India  ink,  collargol)  is  the  liver,  spleen  and  bone 
marrow.  This  was  demonstrated  by  intravenous  injection  of  stained 
fat  suspensions  (lanolin  with  indophenol  —  fat  with  scarlet  R).^ 

In  these  organs  there  are  certain  ceUs  which  take  up  the  fat  par- 
ticles (in  the  liver  the  star  cells  of  von  Kupfer).  It  should  be  again 
emphasized,  that  the  fat  particles  behave  exactly  like  inorganic 
suspensions.  They  behave  like  inorganic  suspensions,  also  as  regards 
rapidity  of  deposition;  (depending  on  the  size  of  the  animal)  after  an 
emulsion  is  poured  into  the  blood  (by  alimentation  or  injection)  one- 
half  hour  to  an  hour  suffices  for  it  to  disappear  from  the  circulation. 

It  follows  from  the  above  that  in  the  deposition  of  fat  there  is  no 
specific  kind  of  fixation;  that  there  is  no  solution  but  a  purely  me- 
chanical retention  of  fat  particles  in  these  storehouses. 

How  the  storage  of  fat  occurs  in  other  organs,  and  how  the  mobili- 
zation of  fat  reserves  must  be  pictured,  whether  as  an  exceptionally 
fine  emulsion  or  a  true  solution,  all  these  are  still  open  questions. 

An  especially  instructive  example  of  research  regarding  an  assimila- 
tion process  is  that  of  WOOD  FORMATION. 

From  a  chemical  standpoint,  the  lignified  cells  of  the  plant  (wood) 
consist  of  protoplasm,  cellulose  and  lignin.  While  cellulose  may  be 
considered  as  a  distinct  substance,  a  highly  polymerized  carbohydrate, 
fittle  has  been  known  regarding  the  constitution  of  "lignin."  Evi- 
dently it  is  a  mixture  of  various  vegetable  gums,  pectins,  lignic  acid, 
albumins,  glucosides,  tannins,  vegetable  coloring  matter,  resins  and 
other  incidental  constituents.  A  substance  in  lignin  which  is  re- 
garded as  characteristic  and  which  stains  with  anilin  salts  and 
phloroglucin  hydrochlorid  was  isolated  by  F.  Czapek  and  identified 
as  an  aromatic  aldehyd. 

Various  theories  have  been  proposed  to  account  for  the  formation 
of  wood;  some  placing  more  stress  on  physiological  cnanges,  and 
others  attempting  to  explain  the  process  on  a  purely  chemical  basis. 
I  can  dismiss  these  investigations  in  view  of  the  fact  that  H.  Wisli- 
CENUS*  has  offered  and  experimentally  established  a  theory  which 
places  the  entire  view  of  this  question  as  well  as  its  experimented 
investigation  upon  a  new  basis. 

He  assumes  that  the  cambial  juice  which  penetrates  the  cambial 
tissue  stored  between  the  wood  and  the  inner  bark  layer  during  the 
summer  vegetative  activity,  contains  crystalloids  (salts,  sugars  and 
plant  acids)  as  well  as  colloids.  These  same  colloidal  constituents 
(formative  substance  or  procambium)  are  all  found  in  the  lignin. 

According  to  H.  Wislicenus,  the  process  of  wood  formation  occurs 
in  three  stages. 

^  For  further  references  see  S.  Bondi  and  A.  Neumann  loc  dt. 


METABOLISM  AND  THE  DISTRIBUTION  OF  MATERIAL     249 

1.  Formation  of  cellulose  hydrogel  in  the  youngest  plant  tissues 
as  a  chemically  indifferent  surface  or  framework.  This  primary 
stage  will  be  explained  more  fully. 

2.  The  colloidal  constituents  of  cambial  juice  become  layered  upon 
the  cellulose  surface  by  adsorption  and  gel-formation,  and  thus  in- 
crease the  thickness  of  the  surfaces. 

3.  Chemical  reactions  occur  between  the  adsorbed  hydrogels  which 
lead  to  lignin  formation. 

The  following  facts  indicate  the  truth  of  these  assumptions: 

(a)  Colloidal  constituents  can  be  extracted  by  adsorption  from  the 
cambial  juice. 

(6)  The  colloids  adsorbable  from  the  cambial  juice  and  the  run- 
ning sap  are  indicative  of  their  hgnin  content,  which  means  their 
wood-forming  properties. 

(c)  The  quantity  of  adsorbable  colloidal  constituents  in  the  cambial 
juice  varies  with  the  season  of  the  year  —  (shown  in  the  annular 
rings  as  early  summer  and  late  summer  wood). 

Certain  trees  give  such  large  quantities  of  spring  sap  on  tapping, 
that  it  is  easy  at  times  to  collect  a  liter  or  more  in  a  day.  In  North 
America  the  sugar  maple  has  this  property.  In  Norway,  Sweden 
and  Russia  people  drink  the  sap  of  the  birch  either  fresh  or  fermented. 
The  trees  are  bored  about  10  cm.  deep,  30  to  40  cm.  above  the  ground 
and  a  glasp  tube  is  inserted  and  sealed  in  with  tree-wax.  The  sap 
drops  through  the  bent  tube  into  a  bottle. 

To  obtain  the  cambial  sap,  trunks  of  birch,  pine  and  gray  ash  are 
sawed  into  pieces,  15  to  20  cm.  long.  The  bark  from  7  to  15  kilos 
of  this  is  taken  and  then  spht  vertically.  The  smooth  inner  layer 
of  the  bark  and  the  outer  cambial  mass  of  the  smooth  surface  of 
wood  are  well  shaved  off  with  glass.  The  shavings  are  placed  in 
from  1  to  2  liters  of  water  and  allowed  to  remain  several  hours,  a  few 
drops  of  thymol  solution  being  added.  The  water  is  poured  off,  the 
residue  squeezed  in  a  fruit  press  and  the  combined  turbid  fluids  are 
filtered. 

The  adsorption  experiments  were  performed  partly  by  shaking 
with  finely  divided  cellulose  (filter  paper)  and  partly  by  siphoning 
through  "washed  clay."  (See  p.  110.)  In  both  cases  the  quantity 
of  material  adsorbed  was  estimated  by  determining  the  weight  of 
the  dried  residue  of  (a)  a  measured  quantity  of  fluid  before  adsorp- 
tion, (6)  the  same  volume  after  adsorption. 

H.  WiSLicENUS  was  able  to  prove  in  the  case  of  the  rising  sap 
obtained  by  tapping  the  hornbeam  and  in  the  cambial  juice  of  the 
birch,  that  the  abstraction  of  colloidal  substances  followed  an  adsorp- 
tion curve,  because  the  more  dilute  the  solutions  the  more  eolloida' 


250  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

substance  relatively  was  extracted  from  them  by  "fibrous  clay"  or 
filter  paper. 

It  appeared  furthermore,  that  the  rising  sap  of  the  birch  contains 
only  a  small  quantity,  from  3.5  to  8.4  per  cent  of  dry  colloid  substance. 
From  this  fact  Wislicenus  concluded  that  in  the  rising  sap  (until 
about  the  opening  of  the  leaves  —  end  of  April)  there  is  no  new  for- 
mation of  colloids,  but  there  occurs  a  dissolving  of  everything  that 
is  soluble  (partial  reversal  of  wood  formation). 

The  cambial  juice  on  the  other  hand  contains,  at  the  time  of  most 
active  wood  formation  (end  of  May  to  end  of  July)  large  quantities 
of  adsorbable  colloids  (24  to  37  per  cent).  Towards  the  end  of  July 
or  beginning  of  August,  when  wood  formation  quickly  ceases,  the 
colloid  content  of  cambial  sap  also  decreases  rapidly. 

p.        (  beginning  of  July 31.1    per  cent  adsorbed  colloids. 

(  beginning  of  August 6.41  per  cent         "  " 

p  ,     (  beginning  of  July 24. 19  per  cent         "  " 

(  beginning  of  August 8.04  per  cent         "  " 

Thus  H.  Wislicenus  has  convincingly  demonstrated  that  wood 
formation  is  a  colloid-chemical  process. 


Enzymes  without  doubt  play  a  most  important  part  in  the  de- 
velopment of  organs.  We  know  that  enzymes  serve  the  body  by 
changing  colloids  into  crystalloids.  They  split  albumin  into  Poly- 
peptids and  amino  acids,  starch  into  saccharids,  etc.  Construction 
or  synthesis  is  also  brought  about  by  enzymes.  Reactions  which  are 
hastened  by  enzymes  are  reversible  and  it  depends  entirely  upon  sur- 
rounding conditions,  whether  the  balance  of  the  process  weighs  more 
in  one  direction  than  in  another.  Thus,  for  instance,  A.  Croft  Hill 
was  the  first  to  show  that  the  same  ferment  which  splits  maltose  into 
glucose  actually  forms  maltose  in  a  concentrated  solution  of  glucose. 
Since  then,  similar  reversals  have  been  frequently  observed:  Potte 
viN  split  fats  by  means  of  pancreatic  lipase  and  with  the  same 
enzyme  he  also  prepared  fats  from  glycerin  and  oleic  acid.  The  well- 
known  cleavage  of  fats  with  the  enzyme  of  the  castor  bean  was  so 
successfully  reversed  by  Welter  that  he  obtained  synthetically 
almost  30  per  cent  of  neutral  fat  with  the  same  enzyme. 

In  general  the  cleavage  process  proceeds  best  in  the  presence  of 
much  water,  whereas  synthesis  is  most  favored  by  the  absence  of 
water.  By  the  swelling  or  shrinking  of  the  colloids  present  during 
the  reaction,  the  organism  is  able  to  permit  the  process  to  proceed 
in  one  or  the  other  direction.     Swelling  and  shrinking  are  in  turn  de- 


METABOLISM  AND   THE  DISTRIBUTION  OF  MATERIAL     251 

pendent  upon  the  formation  and  removal  of  acids  by  oxidative  proc- 
esses. The  concentration  of  the  products  of  a  reaction  in  the  solution 
brings  the  reaction  to  a  standstill.  If  we  are  dealing  with  cleavages 
the  crystalloid  products  may  be  readily  removed  by  diffusion.  This 
does  not  occur  m  the  case  of  synthetic  colloid  products.  Nor  is  it 
so  essential  because  from  the  point  of  view  of  the  law  of  mass  action 
they  are  not  to  be  regarded  as  dissolved. 

We  know  enzymes  to  be  the  excreta  of  the  stomach  and  intestines, 
but  we  also  know  that  cells  themselves  contain  enzymes;  we  may 
mention  the  uricolytic  enzyme  of  the  hver  and  zymase  of  yeast 
which  ferments  sugar. 

We  must  also  recall  that  enzymes  are  the  most  strongly  adsorbed 
of  all  the  colloids  of  the  body,  and  that  the  ability  to  be  adsorbed  is 
largely  dependent  on  the  acid  or  alkaline  character  of  the  medium. 
An  enzyme  may  be  so  fixed  (e.g.,  rennet  by  charcoal)  that  its  very 
existence  is  no  longer  determinable;  a  change  of  reaction  recalls  it 
to  life.  It  may  also  be  released  by  the  approach  of  another  colloid 
(in  our  example  casein)  by  which  it  is  adsorbed  still  more  strongly. 
We  thus  get  an  inkling  of  the  great  importance  enzymes  have  for  the 
life  of  the  cell,  without  as  yet  understanding  the  details. 

The  conditions  governing  dissimilation  are  much  more  readily  un- 
derstood. Through  enzymatic  cleavage  of  colloids,  there  are  formed 
crystalloid  products  which  pass  into  the  circulating  fluids  of  the  or- 
ganism by  diffusion  and  leave  as  excreta,  or,  after  oxidation  to  CO2, 
are  expired. 

We  must  not  conclude  that  in  every  instance  the  entire  colloid 
molecule  breaks  down  into  crystalloid  cleavage  products.  In  this 
way,  by  the  splitting  off  of  individual  "side  chains"  (P.  Ehrlich) 
there  is  permitted  great  variation  in  cell  fife,  which  we  might  assume 
from  our  previous  experiences. 


CHAPTER  XV. 
GROWTH,   METAMORPHOSIS  AND   DEVELOPMENT. 

Growth. 

Of  all  the  problems  in  biology,  one  of  the  most  difficult  and  most 
engrossing  is  the  development  to  constant  type.  From  cells,  which 
externally  can  hardly  be  distinguished,  we  see  develop  a  quail,  an 
oak  tree,  a  butterfly  or  a  man.  In  their  evolution  they  always  pass 
through  the  same  stages  to  the  same  ultimate  forms  which  after  a 
progressive  senescence,  return  to  the  eternal  process  of  evolution. 

If  we  try  to  reduce  these  developmental  processes  to  their  sim- 
plest terms,  we  find  diffusion  and  swelling  phenomena  with  the 
formation  of  precipitate-membranes. 

E.  F.  Runge,  who  discovered  carbohc  acid  in  coal  tar,  and  who 
made  the  first  anilin  color,^  published  a  book  in  1855,  which  is  one  of 
the  most  original  scientific  diversions  I  have  ever  seen.     It  is  called 

"Der  Bildungstrieb  der  Stoffe'' 

(The  formative  instinct  of  matter) 

viewed  in  automatically  developed  figures 

By  Dr.  F.  E.  Runge. 

(Oranienburg.        Printed  by  the  Author.) 

The  book  consists  of  a  collection  of  blotting  paper  leaves,  upon 
which  various  inorganic  salt  solutions  were  dabbed;  they  interacted 
and  gave  colors  by  which  the  most  remarkable  figures  were  produced. 
At  first  glance  these  seem  to  be  lower  forms  of  animal  life,  amebse 
or  rhizopodse,  and  the  collection,  just  as  Häckel's  ''Kunstformen 
der  Natur,"  might  well  serve  as  a  text  for  designers,  because  it 
offers  such  a  multitude  of  suggestions  with  respect  to  color  and 
shape.  All  the  pages  of  the  collection  were  prepared  by  the  author 
himself  (not  printed)  and  are  accompanied  by  a  small  amount  of 
text  which  explains  the  method  of  preparation. 

The  explanation  of  these  creations  is  easy  to  the  author,  who  says 
in  one  of  his  conclusions: 

^  [Chas.  Lowe  is  regarded  as  discoverer  of  phenol  by  the  English,  and  Sir  Wm. 
Henry  Perkin  is  generally  acknowledged,  even  by  Germans,  to  be  the  discoverer 
of  the  first  anilin  color,  mauve.    Tr.] 

252 


GROWTH,  METAMORPHOSIS  AND  DEVELOPMENT         253 

"After  all,  I  believe  I  may  make  the  assertion  that  the  creation 
of  these  pictures  is  due  to  a  new  and  hitherto  unrecognized 
force.  It  has  nothing  in  common  with  magnetism,  electricity 
or  galvanism.  It  is  not  stimulated  or  created  by  any  external 
force  but  is  innate  in  substances  and  becomes  active  when  their 
chemical  affinities  neutralize  themselves ;  that  is,  they  undergo 
selective  attractions  or  repulsions,  and  thus  combine  or  sepa- 
rate. I  call  this  force  '  Formative  Instinct '  and  regard  it  as 
the  prototype  of  the  'vital  force'  of  plants  and  animals." 

Of  course,  no  special  "Force"  need  be  invoked  for  the  explanation 
of  Runge's  pictures.  They  are  the  result  of  very  complicated 
diffusion  and  capillary  phenomena  associated  with  chemical  trans- 
formations. 

What  is  especially  interesting  in  Runge's  pictures,  on  the  one 
hand,  is  the  constancy  of  the  forms  obtained  by  employing  similar 
substances,  and  on  the  other  hand,  the  extraordinary  multiplicity 
brought  about  by  the  diverse  action  of  different  substances. 

If  we  let  a  drop  of  copper  sulphate  solution  fall  on  a  piece  of 
filter  paper  moistened  with  potassium  hydrate,  at  the  surfaces  of 
contact  a  membrane  of  copper  hydroxid  forms,  which  changes 
rapidly  but  always  in  the  same  way.  If  we  always  employ  the  potas- 
sium hydrate  and  copper  sulphate  in  the  same  concentration,  the 
copper  hydroxid  bomidary  will  always  have  the  identical  form, 
provided  the  same  filter  paper  is  used.  A  change  in  the  concen- 
tration of  one  or  the  other  ingredients,  however,  gives  a  membrane 
of  different  shape.  If,  instead  of  copper  sulphate,  we  place  a  drop 
of  copper  nitrate  on  the  paper,  we  obtain  forms  entirely  different, 
and  a  drop  of  nickel  sulphate  changes  the  picture  completely.  We 
thus  see  that  small  variations  in  the  concentration  of  the  solutions 
and  in  their  chemical  composition  possess  numerous  possibilities 
for  the  formation  of  new  shapes. 

In  the  living  organism  variations  in  concentration  perpetually  occur. 
We  know  from  biological  reactions  that  not  only  different  animals, 
as,  for  instance,  sheep  and  lions,  have  chernically  different  tissues,  but 
that  even§the  ass  and  the  horse,  and  indeed  different  races  of  men, 
may  be  chemically  differentiated;  consequently  the  second  condition 
for  variation  in  form  is  also  given,  namely,  the  difference  in  chemical 
composition.  The  processes  of  the  body  (organism)  are  regulated  by 
its  colloidal  state,  and  this  very  colloidal  state  also  permits  the  reteji- 
tion  of  shapes. 

If  we  seek  to  leave  this  far  too  general  point  of  view  and  study 
details  of  the  question  more  closely,  we  encounter  almost  insur- 
mountable difficulties. 

If  a  small  lump  of  copper  sulphate  is  thro^vn  into  a  dilute  solution 


254  COLLOIDS  IN  BIOLOGY  AND-  MEDICINE 

of  potassium  ferrocyanid,  there  will  soon  develop  a  brown  envelope 
which  throws  out  upward-growing  runners,  and  in  half  an  hour's 
time  the  fluid  is  filled  with  figures  which  vividly  recall  both  the 
shape  and  the  color  of  seaweed;  if  a  small  amount  (0.5  per  cent)  of 
gelatin  has  been  added  to  the  water,  the  figures  have  some  stability. 
Their  development  is  easily  explained:  the  copper  sulphate  dis- 
solves and  immediately  forms  a  semipermeable  membrane  of  copper 
ferrocyanid,  through  which  no  copper  sulphate  can  escape,  but  water 
may  enter.  Since  a  concentrated  solution  of  copper  sulphate  is 
formed  within,  water  will  be  absorbed  until  the  membrane  bursts^ 
whereupon  the  copper  sulphate  solution  is  brought  into  contact 
with  the  potassium  ferrocyanid  again  and  forms  a  new  pellicle  of 
copper  ferrocyanid,  and  thus  the  process  goes  on. 

Stephane  Leduc  has  studied  these  figures  most  industriously 
and  has  discussed  their  significance  in  numerous  publications.^ 

Some  of  his  directions  are  here  given:  Prepare  granules  of  1  part 
sugar  and  1  or  2  parts  copper  sulphate.  This  is  scattered  in  a  fluid 
consisting  of  100  parts  of  water  and  from  10  to  20  parts  10  per  cent 
gelatin,  5  to  10  parts  saturated  potassium  ferrocyanid  solution  and 
5  to  10  parts  saturated  sodium  chlorid  solution,  which  mixture  has 
been  heated  to  40  degrees  Centigrade.  In  this  way  we  obtain  figures 
Uke  that  in  Fig.  41,  which  may  attain  a  height  of  40  cm.  The  gelatin 
is  solidified  by  cooling  and  the  figures  may  be  preserved.  Other  fig- 
ures are  obtained  by  throwing  granules  of  fused  calcium  chlorid  or 
barium  chlorid  into  a  concentrated  solution  of  soda. 

Another  recipe  is: 

Water 1  liter 

33  per  cent  potassium  water  glass  solution 60  gm. 

Saturated  soda  solution 60  gm. 

Saturated  sodium  phosphate  solution 60  gm. 

Beautiful  branching  figures  are  given  by  scattering  calcium  chlorid 
granules  in  this  mixture. 

The  more  concentrated  the  solutions,  the  more  rapid  is  the  growth 
and  the  more  branching  and  delicate  are  the  shapes.  If  the  outer 
water  is  diluted  while  the  growth  is  in  progress,  we  may  produce 
figures  with  stems  and  tops,  like  fungi  (mushrooms  and  toadstools, 
etc.).  These  figures  react  to  small  changes  in  osmotic  pressures  by 
changing  their  shape. 

^  I  shall  mention  only  his  latest  pubhcations:  St.  Leduc,  Biochem.  Ztschr. 
(Festband  f.  H.  J.  Hamburger),  1908,  280  u.  ff.  —  Les  croissances  osmotiques  et 
I'origine  des  etres  vivants  (Bar-le-Duc,  1909).  Les  bases  physiques  de  la  vie  et 
la  biogenese  (Presse  Medicale,  7,  12,  1909).  Theorie  physico-chimique  de  la  vie 
(Paris,  1910),  La  dynamique  de  la  vie  (A.  Poinat,  Paris,  1913) ;  and  many  others. 


GROWTH,   METAMORPHOSIS  AND  DEVELOPMENT         255 

St.  Leduc  calculated  that  a  granule  of  potassium  ferrocyanid 
acquired  150  times  its  original  weight  when  it  grew,  and  that  calcium 
structures  acquired  many  hundred  times  their  original  weight. 

The  internal  structure,  also,  has  some  similarity  to  natural  forms. 
They  have  a  cellular  structure  as  St.  Leduc  showed  by  microphoto- 


FiG.  41.    Artificially  prepared  osmotic  seaweed.     (Made  by  St.  Leduc.) 


graphs,  Fig.  42,  and  as  could  be  inferred  from  the  way  in  which  they 
are  formed.  The  solution,  e.g.,  of  copper  sulphate,  which  is  sur- 
rounded by  a  membrane  of  copper  ferrocyanid,  imbibes  water  until 
the  membrane  bursts,  and  some  copper  sulphate  solution  is  exuded, 
which  straightway  surrounds  itself  with  a  film  of  copper  ferrocyanid, 
and  thus  a  new  cell  is  formed.  The  process  repeats  itself  and  one 
cell  is  added  to  another. 

It  cannot  be  denied  that  the  pictures  reproduced  have  a  marked 
external  resemblance  to  natural  algse,  fungi,  etc.,  and  that  it  is 


256 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


possible  to  imitate  nucleus  and  cell  division,  growth  and  all  sorts  of 
phenomena,  and  that  even  their  internal  structure  is  in  many  re- 
spects suggestive  of  a  cellular  structure.  Doubtless  structures  do 
occur  in  nature  which  develop  in  the  same  way  as  these  artificial 
osmotic  products.     In  fruit  wines  after  fermentation,  H.  Miller- 


Fig.  42.     Microphotographs  of  osmotic  structures,  showing  a  cellular  structure. 
Magnified  X  60.     (From  St.  Leduc.) 


Thürgaü*  found  vesicles  which  were  filled  with  bacteria  (Fig.  43) 
on  the  yeast  sediment.  These  "bacteria  vesicles"  were  developed 
because  the  colloid  substances  eliminated  by  the  bacteria,  in  con- 
junction with  the  fruit  wine,  which  contains  tannin,  form  a  semi- 
permeable membrane,  a  vesicle,  that  grows  and  sends  out  tubules. 

Is  there  really  an  analogy  in  the  development  of  these  structures  to 
the  development  of  natural  organisms?  ^  The  fact  that  there  is  not  the 
slightest  chemical  resemblance  to  organisms  may  be  completely 
disregarded  since  St.  Leduc  and  all  who  share  his  views  speak  only 
of  the  similarity  of  the  physical  force  at  work  in  both. 

1  W.  Roux  has  treated  the  entire  question  in  a  very  instructive  essay  on  the 
"Angebliche  Kunstliche  Erzeugung  von  Lebewesen"  in  the  "Umschau"  (Frank- 
fort a.  M.),  1906,  Nr.  8. 


GROWTH,   METAMOliPHOSIS  AND  DEVELOPMENT         257 

It  is  a  detriment  to  the  scientific  treatment  of  tlie  entire  question 
that  the  outward  resemblance  (form  and  color)  is  so  strikingly  hke 
the  natural  structures.  Involuntarily,  we  are  reminded  of  wax 
figures  which  move  their  arais  and  legs.  Though  the  internal  struc- 
ture has  some  slight  resemblance  to  some  natural  organisms,  the 
analogy  completely  fails  if  a\  e  consider  such  de- 
tails as  cell  division  and  cell  multiplication.  The 
figures  of  St.  Leduc  absorb  no  nourishment, 
other  than  water;  their  increased  weight,  as  far 
as  solid  substances  go,  consists  only  in  pellicle 
formation,  and  in  spite  of  a  shape  suggestive  of 
higher  organisms,  they  have  no  differentiated 
internal  structure,  and  the  cell  division  has  not 
the  remotest  resemblance  to  natural  cell  division, 
but  occurs  intermittently  by  bursting,  etc.  We 
might,  if  it  did  not  seem  fruitless,  multiply  the  Yig  43  Vesicles  of 
dissimilarities  indefinitely  and  we  might  show  Bacterium  mani- 
that  metabolism  and  the  development  of  germ  topoem  from  a 
cells  is  out  of  the  question  in  such  formations.  P^^  culture  in 
■vrr  i        X  1      1      •       4.1,  £      u         sterile  pear  juice. 

We  must  not   overlook,  m  the  presence   ot   ail       r^,         •  i  i.     j 

.  '  1  1       •      1     r.  1  he  vesicle  has  de- 

these    dissimilarities,    that    the    physical    forces       veloped     a    long 

which   produce   these   inorganic   formations   are       transparent  tube, 
the  same  as  those  ivhich  produce  the  growth  and       Magnification 

configuration   of  organized   material:    membranes,       200:1.    (After  H. 

,.  j--ff      •  MüUer-Thm-gau.) 

osmotic  pressure,  dijjusion.  ^     ' 

In  one  point,  at  least,  Leduc's  analogies  fail  completely:  except- 
ing the  membranes,  they  are  devoid  of  colloid  material.  We  have 
already  seen  that  swelling  frequently  replaces  osmotic  pressure. 
If  we  could  imagine  the  crystalloid  material  of  St.  Leduc 
replaced  by  colloids  capable  of  swelling,  we  would  have  the  essen- 
tial physical  and  chemical  conditions  for  growth  and  structure  of 
organisms. 

A  serious  study  of  these  problems,  one  which  extends  beyond  ex- 
ternal resemblances,  is  still  in  its  earliest  beginnings;  see  R.  E.  Liese- 
gang, Nachahmung  von  Lehensvorgängen.  Formations  resembhng 
the  creations  of  St.  Leduc  have  been  independent^  noticed  by 
others.  B.  D.  Uhlenhuth*  produced  beautiful  growths  b}^  putting 
iron  objects  into  antiformin.  Antiformin  is  a  mixture  of  sodium 
hypochlorite  with  sodium  hydrate.  The  formations  consist  of 
iron  oxid,  and  their  development  is  easily  understood  from  the  ex- 
planations that  have  been  already  given.  Since  a  small  amount  of  a 
water-soluble  iron  salt  must  be  formed  first  by  the  action  of  the 
hypochlorite  of  soda  on  the  iron,  the  growth  is  slower,  the  figures 


258  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

are  more  beautiful  and  possibly  more  natural.     In  two  weeks  the 
structures  attain  a  height  of  from  5  to  10  cm. 

The  growths  described  above  offer  analogies  to  organisms  which 
are  completely  surrounded  by  water  only,  yet  there  are  those  which 
offer  a  resemblance  to  the  growth  of  terrestrial  plants.  We  might 
mention  the  ''blossoms"  of  many  crystalloid  substances,  especially 
the  ammonia  salts.  H.  Wislicenus*^  has  thoroughly  studied  the 
growth  and  structure  of  fibrous  alumina.  If  granules  of  aluminium 
which  have  been  activated  by  contact  with  traces  of  mercury  subli- 
mate, for  example,  are  permitted  to  he  in  a  moist  place,  very  soon 


Fig.  44.    Fibers  of  fibrous  alumina,  magnified  X  40.     (From  H.  Wislicenus.) 


fibrous  structures  grow  from  the  metal,  which  in  a  few  hours  may 
reach  more  than  1  cm.  in  length.  Under  the  influence  of  the  mercury 
as  a  catalyzer,  aluminium  oxid  is  formed  from  the  aluminium  accord- 
ing to  the  formula : 

Al  +  3  H2O  =  Al  (0H)3  +  3  H. 

In  this  instance  it  is  not  the  osmotic  pressure  of  entering  water 
which  bursts  the  films,  thus  bringing  a  fresh  metal  surface  to  the 
reaction,  but  the  pressure  of  the  hydrogen  gas. 

Though  in  this  case,  as  well,  there  are  many  gaps  in  the  resemblance 
to  natural  organisms,  nevertheless  the  fibers  formed  show  certain 
resemblances  to  real  fibers  (see  Figs.  44,  45). 


GROWTH,   METAMORPHOSIS  AND  DEVELOPMENT         259 

In  the  first  place  they  resemljle  most  organic  fibers  by  being 
doubly  refractive  (L.  Jost*)  even  though  this  doul^le  refraction  is 
caused  by  different  conditions  than  in  organized  structures.  In 
contradistinction  to  natural  fibers,  the  substance  is  isotropic;  it  is 
only  its  lamellated  structure  which  produces  that  type  of  double  re- 
fraction (H.  Ambronn*)  which  is  found  in  the  siliceous  envelopes  of 


Pig.  45.    Piece  of  doublj'-  diffracting  fibrous  alumina.    Magnified  X  440. 
(After  H.  Wislicenus.) 

diatoms  (e.g.,  pleurosigma  and  amphipleura)  and  probably  also  in 
tabasheer,  the  colloidal  silicic  acid  which  occurs  in  the  internodes  of 
some  species  of  bamboo. 

The  Genesis  of  Structures. 

What  phase  is  for  the  physical  chemist,  cells  and  tissues  are  for 
the  biologist.  Like  phases,  cells  and  tissues  are  "portions  of  a 
structure  separated  from  one  another  by  physical  interfaces"  (Wil- 
helm Ostwald's  definition  of  phase).  The  interface  maj^  consist 
of  an  invisible  transition  layer  (see  p.  280).  The  interface  is  most 
evident  when  it  consists  of  a  visible  membrane.  Such  a  membrane 
whether  visible  or  invisible  is  always  a  structure  poor  in  water.     At 


260  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

the  interface  against  air  it  may  be  created  by  desiccation.  Within 
the  organism,  we  must  assmne  that  membranes  develop  similar  to 
chemical  precipitation-membranes.  If  we  add  silver  nitrate  to  a 
solution  of  common  salt,  a  precipitate  of  silver  chlorid  is  formed. 
If  we  permit  common  salt  and  silver  nitrate  to  diffuse  together  in  a 
jelly,  at  the  point  of  contact  a  membrane  of  AgCl  develops.  We  con- 
sidered the  results  of  this  more  thoroughly  in  the  introduction  to 
this  chapter.  It  is  now,  therefore,  merely  necessary  to  recall  that  not 
only  may  crystalloids  form  such  membranes  in  a  jelly,  but  that  with 
albuminous  material  H.  Bechhold*^  produced  such  membranes  in  jel- 
lies (phosphoric  acid,  goat  serum  and  goat-rabbit  serum).  Theoreti- 
cally, therefore,  the  development  of  membranes  offers  no  difficulties. 

The  development  of  a  precipitate  in  a  jelly  gives  a  certain  direction 
to  the  further  evolution  of  the  process.  The  sense  in  which  this  is 
intended  will  be  elucidated  by  some  examples;  by  membrane  forma- 
tion, to  begin  with.  Substances  which  form  no  membrane  by  precipi- 
tation, diffuse  together  unhindered,  and  in  time  become  completely 
mixed.  If  a  semipermeable  membrane  has  formed,  it  behaves  like  a 
solid  wall  that  arrests  any  further  mixture.  If  two  solutions  of  equal 
osmotic  pressure  diffuse  together  in  a  jelly  till  they  form  a  permeable 
membrane,  no  matter  how  thin,  e.g.,  sodium  chlorid  and  silver  nitrate, 
diffusion  ceases  as  soon  as  the  membrane  has  a  very  slight  thickness. 
If,  however,  the  osmotic  pressure  on  one  side  is  greater,  the  mem- 
brane continues  to  grow  until  the  osmotic  pressure  is  equal  on  both 
sides  (N.  Pringsheim,*  H.  Bechhold  and  J.  Ziegler*^). 

The  phenomena  are  exceedingly  interesting  when  precipitates  de- 
velop simultaneously  in  several  places.  These  phenomena  have  been 
studied  by  R.  Liesegang.*^  If  we  place  on  a  plate  which  is  covered 
with  sodium  chlorid  jelly,  a  drop  of  silver  nitrate,  there  forms  a 
disc-shaped  precipitate  of  silver  chlorid,  whose  circumference  in- 
creases equally  in  all  directions  (a  circle)  according  as  the  silver 
nitrate  diffuses  into  the  sodium  chlorid  jelly.  If,  however,  two 
drops  are  placed  on  the  sodium  chlorid  jelly  several  centimeters 
apart,  there  develops  a  picture  like  Fig.  46;  the  two  silver  chlorid 
precipitates  grow  towards  each  other,  that  is,  an  ''apparent  chemical 
attraction"  is  observed.  The  reason  for  this  is  as  follows:  im- 
mediately upon  applying  the  silver  nitrate,  the  jelly  loses  sodium 
chlorid  because  of  the  precipitation  of  AgCl,  and  this  causes  a 
movement  in  the  entire  mass  of  sodium  chlorid:  the  spot  where  the 
precipitate  forms  is  deprived  of  chlorin  ions,  which  then  diffuse 
in  afresh  from  the  periphery.  If  two  neighboring  drops  of  silver 
nitrate  have  been  placed  on  the  sodium  chlorid  jelly,  there  forms  be- 
tween them  a  region  poor  in  chlorin,  which  thus  permits  a  more  rapid 


Fig.    51.     Laminated     urinary    calculus 
Fig.  50.     Liesegang 's  rings.      (From  (urates).     (Drawing    by    H.    Schadde; 

R.  Liesegang.)  horn  von  Frisch  and  E.  Zuckerkandl.) 


Fig.  53.  Oil-droplets  super-saturated  with  Cho- 
lesterin. Crystalline  separation  of  Cholesterin 
may  be  recognized  in  several  droplets. 


Fig.  52.  Primitive  gall-stone 
pattern.  (Myelin  clump.) 
Magnified  X  62. 


Fig.  54.  Laminated 
calcium  bilirubin 
stone. 


PLATE  II. 


GROWTH,  METAMORPHOSIS  AND  DEVELOPMENT         261 

advance  of  the  silver  nitrate.  What  is  shown  here  in  the  case  of  the 
silver  chlorid  holds  for  every  other  precipitate,  and  for  every  osmotic 
disturbance,  provided  only  that  dif- 
fusible substances  are  present  in  a 
jelly. 

If  such  disturbances  (precipi- 
tates, membranes)  occur  simultane- 
ously in  various  places,  it  is  possible 
«       .,  ,  T     J.    1  r:  J.       Fig.  46.     Apparent  chemical  attrac- 

for  the  most  complicated  fagures  to  ^.         ,-r,  t-  ^ 

^  °  tion.     (R.  Liesegang.) 

form. 

In  addition,  we  must  consider  the  changes  which  occur  in  the  ordi- 
nary course  of  swelling  and  shrinking  of  the  colloidal  material. 

As  far  as  I  know,  it  has  hitherto  not  been  possible  to  explain 
such  a  phenomenon  in  vivo.  I  wish  to  refer  to  only  one  analogy: 
C.  U.  Ariens-Kappers*  described  as  neurobiotaxis  a  phenomenon  of 
nerve  fibers:  if  two  nerve  cells,  a  certain  distance  apart,  are  injured 
simultaneously  or  in  close  succession,  the  growth  of  the  chief  dendron 
of  both  injured  ganglion  cells  occurs  in  the  direction  of  the  other 
stimulated  or  injured  cell.  We  thus  have  a  growth  towards  each 
other,  analogous  to  the  apparent  chemical  attraction  just  described. 

[C.  A.  Elsberg  concludes  from  his  experiments  that  hyperneuroti- 
zation  of  a  normal  muscle  is  impossible.  A  normal  muscle  cannot  be 
made  to  take  on  additional  nerve  supply.  The  implanted  nerve  can- 
not make  neuro-motor  connections.  If  the  muscle  is  permanently  sep- 
arated from  its  original  nerve,  the  implanted  nerve  will  then  establish 
such  connections.     Science  N.  S.,  Vol.  XLV,  p.  319  et.  seq.     Tr.] 

Naturally,  cells  mutually  modify  each  other's  shape.  A  structure 
which  would  develop  spherically  if  uninfluenced,  under  the  pressure  of 
neighboring  cells  acquires  a  reticulated,  fibrous  or  pavement  shape. 

Layered  Structures. 

We  saw  that  if  two  solutions  which  form  a  precipitate  meet  in  a 
jelly,  a  precipitation  membrane  develops  at  the  point  of  contact. 
Provided  this  is  sufficiently  permeable  and  one  solution  has  a  higher 
osmotic  pressure,  the  membrane  continues  to  grow  uninterruptedly, 
becoming  constantly  thicker  until  the  osmotic  pressure  is  the  same 
on  both  sides.  In  1898,  R.  E.  Liesegang*^  pubhshed  an  observation 
which  does  not  accord  with  the  continuous  growth  mentioned  above. 

If,  for  instance,  ammonium  bichromate  is  dissolved  in  melted 
gelatin,  which  is  then  solidified  in  shallow  dishes,  and  upon  it  a  drop 
of  silver  nitrate  is  placed,  then  there  does  not  develop  upon  diffusion  a 
constantly  thicker  precipitation  membrane  of  silver  Chromate  but  con- 
centric rings  called  Liesegang's  rings  (see  Plate  II,  Fig.  50).     The 


262 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


experiment  may  be  performed  by  solidifying  the  ammonium  bichro- 
mate gelatin  in  a  test  tube  and  layering  some  silver  nitrate  over  it. 
We  thus  get  instead  of  rings  true  precipitation  membranes,  which 
are  separated  from  one  another  by  layers  containing  no  silver  Chro- 
mate (see  Fig.  47).  Subsequently,  Wilhelm  Ostwald, *2  J.  Haus- 
mann,* H.  W.  MoESE  and  G.  W.  Pierce,*  H.  Bechhold*^  and 
E.  Hatschek  studied  the  development  of  these  rings.  It  may  be 
assumed  as  a  result  of  these  investigations  that  the  formation  of 
such  layers  is  the  result  of  a  very  complicated  combination  of  events 
whose  further  elucidation  at  this  point  would 
carry  us  too  far  afield. 

It  should  be  definitely  stated  that  the  de- 
velopment of  rhythmic  structures  is  in  no  way 
dependent  on  the  interdiffusion  of  two  solutions. 
Similar  structures  may  be  produced  in  jellies  also 
by  crystallization  (e.g.,  tri-sodium  phosphate)  or 
by  freezing  water. 

The  ring  formation  occurs  especially  when 
ammonium  Chromate  and  silver  nitrate  come  in 
contact;  at  times  there  may  be  produced  as 
many  as  twenty  or  more  parallel  membranes, 
which,  according  to  the  concentration  of  the 
solution,  may  be  separated  from  a  fraction  of  a 
millimeter  up  to  1/2  centimeter.  They  have  also 
been  produced  by  numerous  other  precipitation 
reactions. 

H.  Bechhold  prepared  similar  stratified  mem- 
branes with  organic  material.  This  occurs  readily 
if  serum  mixed  with  gelatin  is  permitted  to  solidify 
in  a  test  tube  and  metaphosphoric  acid  is  layered 
over  it.  The  number  and  beauty  of  the  mem- 
branes depend  very  much  on  the  relative  con- 
centrations of  the  solutions  employed.  Most 
advantageous  is  a  mixture  of  2.5  per  cent  serum 
and  of  5  per  cent  gelatin,  upon  which  is  placed 
2  per  cent  metaphosphoric  acid;  this  gives  as  many  as  five  concentric 
rings.  The  author  obtained  two  parallel  membranes  by  the  diffusion 
of  goat  serum  into  gelatin,  containing  goat-rabbit  serum. 

It  is  evident  in  the  formation  of  this  kind  of  layered  membranes 
that  the  phenomenon  noted  on  page  85  et  seq.  plays  an  important  role: 
colloids  only  precipitate  in  definite  mixture  relations,  and  solution  oc- 
curs in  the  presence  of  an  excess  of  either  one.  This  phenomenon  can 
be  followed  visually  in  the  above-described  membrane  formation. 


jaty-^- 


Fig.  47.  Stratifica- 
tions in  a  test  tube. 
(F.  Stoffel.) 


GROWTH,   METAMORPHOSIS  AND  DEVELOPMENT         263 

It  is  easy  to  imagine  that  layered  membranes  develop  by  the  re- 
moval of  a  substance  which  holds  another  substance  in  solution.  I 
have  experimented  with  this  end  in  view  by  dissolving  globulin  in 
gelatin  containing  sodium  chlorid  and  layering  water  over  it;  when 
the  sodium  chlorid  diffused  away,  then  the  globulin  precipitated 
out.  As  a  matter  of  fact  no  layered  structures  developed  in  the 
gelatin  but  only  turbidities  uniformly  distributed.  This  does  not 
by  any  means  mean  that,  with  a  different  arrangement  of  the  ex- 
periment, regular  layered  membranes  might  not  be  obtained. 

In  organisms  we  frequently  encounter  stratified  structures  which, 
in  most  cases,  occur  as  the  result  of  rhythmic  deposition.  The  an- 
nual rings  of  trees,  the  various  layers  in  the  otoliths  of  young  and  of 
old  fishes  cannot  be  explained  in  any  other  way  than  that  periods  of 


Fig.  48.     Starch  granules.     (Kilnitz-Gerloff.) 

rest  follow  periods  of  strong  accretion.  In  contrast  to  these  "external 
rhythms"  which  obviously  are  induced  by  changing  conditions  affect- 
ing an  organ,  there  are  also  layered  structures  with  "internal  rhythms  " 
which  suggest  Liesegang's  rings  As  such,  we  must  regard  starch 
grains  (Fig.  48),  the  sihcious,  sponginous  and  calcareous  structure  of 
sponges,  and  the  perforated  calcareous  shells  of  the  foraminifera  and 
many  fish  scales.  R.  Liesegang*^  mentions  in  addition  the  con- 
centric lamellae  about  the  Haversian  canals  in  the  bones  of  verte- 
brates, the  rods  of  the  retina  and  the  spiral  cross  strise  of  muscle 
fibers.  W.  Gebhardt  compares  the  rhythmic  markings  on  butterflies' 
wings  to  Liesegang  structures.  The  coarse  layers  of  the  otoliths 
of  fishes,  the  annual  rings  of  trees,  the  concentric  structure  of  pearls 
are  penetrated  by  still  finer  layers,  whose  formation  R.  Liesegang 
thinks  is  analogous  to  the  formation  of  the  rings  he  described. 


264  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

E.  Küster  has  called  attention  to  numerous  rythmic  phenomena 
among  plants  in  which  the  influence  of  external  forces  cannot  be 
repognized  and  which  consequently  he  attributes  to  Liesegang's 
law.  Among  others  we  may  mention  the  thickening  of  membranes 
in  vessels  and  trachese,  the  bands  in  striated  portions  of  plants  (herbs, 
pinus  Thunbergii  Pari.),  and  the  rhythmic  changes  in  many  blos- 
soms. Doubtless  the  number  of  instances  where  we  may  entertain 
the  idea  of  internal  rhythms  may  be  increased  at  will  and  it  will  be 
the  task  of  future  investigators  to  determine  the  essential  causes. 

A  valuable  contribution  in  this  connection  was  made  by  M.  Munk 
who  studied  the  formation  of  fairy  rings.  When  moulds  are  grown 
on  bread,  nutrient  agar,  etc.,  we  frequently  observe  growth  in  con- 
centric rings  which  suggest  Liesegang's  layers  and  are  popularly 
called  fairy  rings. 

M.  Munk  demonstrated  that  the  accumulation  of  metabolic 
products  interfered  with  growth,  causing  a  zone  where  there  was  but 
little  mould.  In  the  case  of  some  strong  acid  producing  moulds 
the  distance  between  the  individual  rings  may  be  regulated  by  the 
addition  of  alkali  to  the  nutrient  medium.  If  litmus  agar  is  used  the 
blue  and  red  rings  make  visible  the  cause  of  the  ring  formation. 

It  is  interesting  that  layered  structures  in  the  ends  of  the  periph- 
eral nerves,  which  were  looked  upon  as  real,  have  been  proved  to 
be  artefacts.  Golgi  stained  nerves  by  saturating  them  with  potas- 
sium bichromate  and  then  treating  them  with  silver  nitrate.  He 
obtained  stratified  structures  whose  appearance,  as  H.  Rabl  showed, 
changed  with  the  concentration  of  the  solutions,  and  they  could  be 
nothing  other  than  Liesegang's  rings. 

Biological  Growth. 

The  fertilization  of  the  egg  is  evidently  the  cause  of  the  powerful 
swelling  processes,  which  are  possibly  induced  by  the  formation  of 
acids.  According  to  Jacques  Loeb,*^  oxidation  processes  accompany 
the  development  of  the  egg  (whether  fertilized  or  parthogenetic) ; 
without  oxygen  no  development  of  the  ovum  occurs.  The  increase 
in  the  volume  of  the  ovum  of  Echinoderma,  until  it  reaches  the 
pluteus  stage,  is  entirely  conditioned  by  the  absorption  of  water 
(C.  Herbst*).  Before  the  larvae  reach  the  pluteus  stage  they  can- 
not a.ssimilate  any  organic  nourishment.  Davenport*  in  the  case  of 
frog  embryos  has  shown  that  their  dried  weight  remains  the  same 
or  diminishes  till  the  moment  when  they  commence  to  eat.  Their 
water  content  on  the  other  hand  was  enormously  increased.  This 
absorption  of  water  is  not  due  to  an  increase  in  the  osmotic  pressure^ 


GROWTH,   METAMORPHOSIS  AND  DEVELOPMENT         265 


since  the  fertilized  and  the  nonfertihzed  frog's  egg  shows,  according 
to  L.  Backmann  and  J.  Rünnstrom,*  only  1/10  of  the  osmotic 
pressure  of  the  egg  in  the  ovary,  or  of  the  adult  frog.  In  the  course 
of  development,  the  osmotic  pressure  increases  so  that  in  the  tadpole 
of  25  or  30  days,  the  osmotic  pressure  is  almost  the  same  as  in  the 
metamorphosed  animal.  L.  Backmann  and  J.  Rünnstrom  agree 
that  the  decrease  in  osmotic  pressure  is  due  to  the  fertilization,  which 
results  in  a  gel  formation  by  means  of  which  crystalloids  are  adsorbed. 
After  a  certain  time,  which  varies  for  different  animals,  but  not 
as  yet  definitely  established  for  individual  ones,  a  shrinking  begins 
again,  as  may  be  seen  from  the  following  data,  taken  in  part  from  the 
tables  of  H.  Gerhartz*^: 

Man. 


Water,  per  cent 

94.0 

90.0 

86.0 

83.3 

71.7 

67.6 

66.0 

Dry  substances, 
per  cent. 


3d  fetal  month 

6th  fetal  month  (Rubner) 
7th  fetal  month  (Rubner) 
8th  fetal  month  (Rubner) 
Newborn  (Camerer,  Jr. ) .  . 

Adult  (Moleschott) 

Adult  (Bouchard) 


Fetus  (I  inch  long)  (A.  v.  Bezold) . 

Newborn  (A.  v.  Bezold) 

8  days  old  (A.  v.  Bezold) 

Full  grown  (A.  v.  Bezold) 


87.2 
82.8 
76.8 
73.3 


6.0 
9.7 
14.0 
16.7 
28.3 
32.4 
34.0 


Dog. 

6  days  old  (Gerhartz) 

80.3 
77.0 

19.7 

15  days  old  (Gerhartz) 

23.0 

Sheep. 

6  months  old  (Lawes  and  Gilbert) 

47.8 
43.4 

52.2 

15  months  old 

56.6 

Mouse. 

12.8 
17.2 
23.2 

28.7 


Chicken  Embryo  (without  yolk). 


7th  day  (L.  v.  Liebermann) , 
14th  day  (L.  v.  Liebermann) . 
21st  day  (L.  v.  Liebermann) 


7.2 
12.7 
19.65 


What  constituents  are  especially  deprived  of  water  cannot  be 
properly  determined  from  the  limited  material  at  hand,  yet,  accord- 


266  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

ing  to  H.  Gerhartz,  there  seems  to  be  a  very  great  shrinking  even 
of  the  albumin.  For  man  he  calculates  the  proportion  of  albumin 
to  water  to  be: 

Newborn 1  albumia,  5.6  water 

Adult 1  albumin,  4.3  water 

J.  A.  KuBowiTSCH  has  shown  that  the  water  content  of  a  mam- 
malian embryo's  muscle  sinks  from  99.4  per  cent  to  8  per  cent  at  the 
termination  of  fetal  life  and  finally  to  75-80  per  cent  in  adults. 
According  to  L.  B.  Mendel  and  Leavenworth,  pig's  hver  has  a 
quite  constant  water  content  of  about  80  per  cent  during  fetal  life 
but  diminishes  to  67.3  per  cent  in  the  adult. 

From  this  we  understand  that  in  the  earliest  stages,  growth  only 
results  by  means  of  the  water  taken  up  through  swelling,  though  a 
time  comes  when  growth  is  induced  by  the  entrance  of  solid  sub- 
stances, by  assimilation.  This  assimilated  substance  meanwhile 
binds  less  water;  with  further  growth  there  is  associated  a  relative 
shrinking  which  after  reaching  its  maximum  (growth)  passes  with 
further  age  into  an  absolute  shrinking. 

According  to  Mühlmann,  aging  of  different  organs  does  not 
proceed  equally.  The  weight  of  human  intestines  increases  up  to 
the  fiftieth  year,  but  the  heart  and  lungs  never  cease  gaining  weight; 
the  brain,  on  the  other  hand,  has  achieved  its  maximum  weight  at 
about  the  end  of  the  second  decade  and  from  then  on  it  gradually 
declines.  The  brain  also  shows  definite  microscopic  aging  phenom- 
ena, even  in  the  earliest  years.  Lipoid  pigment  granules  appear 
in  the  nerve  cells  which  continually  increase  and  at  an  advanced  age 
fill  the  entire  cell  According  to  Marinesco  it  is  much  easier  to 
destroy  with  solvents  suspensions  of  ganglion  cells  of  a  newborn 
puppy  than  an  old  dog.  As  the  result  of  his  studies  of  pigment 
granules  in  nerve  cells  he  also  arrives  at  the  conclusion  that  aging  is 
due  to  the  coagulation  of  physiological  elements,  a  diminution  of 
surface  tension,  such  as  we  know  occurs  in  the  aging  of  colloids 
(see  p.  73). 

H.  Schade  determined  that  the  subcutaneous  connective  tissue  dis- 
solved much  more  rapidly  in  NaOH  when  it  was  derived  from  a  month 
old  child  than  when  it  was  taken  from  a  thirty-two  year  old  woman. 

F.  Tangl*  is  of  the  opinion  that  the  shrinking  of  the  animal 
organism  during  embryonal  development  is  a  duplication  of  the 
same  phylogenetic  process  and  shows  by  numerous  tables  that  the 
lower  invertebrates,  even  those  which  do  not  live  in  water,  are 
usually  more  rich  in  water  than  the  higher  vertebrates. 

By  what  chemism  increase  of  water  and  substance  are  condi- 


GROWTH,   METAMORPHOSIS  AND  DEVELOPMENT         2()7 

tioned,  how  cell  dividon  results,  what  are  the  relations  between 
nucleus  and  cell  protoplasm,  are  questions  which  are  not  yet  ripe 
for  colloid  research. 

To  be  sure  we  know  that  such  swelling  and  shrinking  processes 
occur,  not  only  for  the  entire  cell,  but  also  in  the  nucleus.  Before 
each  cell  division  the  nucleus  swells  up  very  much,  and  after  division 
shrinks  again,  decidedly. 

BoROWiKOW*  made  interesting  observations  on  plant  growth.  All 
who  are  familiar  with  plants  know  that  even  in  summer,  periods  of 
apparent  rest  alternate  with  periods  of  active  growth  (sprouting). 
The  latter  phase  is  associated  with  considerable  entrance  of  water. 
It  is  impossible  to  explain  this  inhibition  of  water  by  osmotic  forces 
since  the  increased  rate  of  growth  was  usually  associated  with  a 
diminution  in  the  concentration  of  the  cell  juices  instead  of  the 
reverse,  and  a  growing  plant  absorbed  unequal  quantities  of  water 
from  solutions  osmotically  identical.  There  was,  on  the  contrary, 
some  evidence  that  sweUing  processes  were  active.  Martin  H. 
Fischer  had  already  called  attention  to  the  fact  that  the  tips  of  buds 
are  always  acid  in  reaction.  It  was  quite  natural,  therefore,  to  test 
the  influence  of  acids,  bases  and  salts  on  the  sprouting  of  plants  and 
to  compare  it  with  the  swelhng  of  colloids.  For  this  purpose  Boro- 
wiKOW  placed  six-day  old  sunflower  seedlings  (HeHanthus  ammus) 
in  sieves  and  dipped  them  in  various  solutions,  using  distilled  water 
as  a  control. 

Dilute  acids  (1/100  normal)  accelerated  growth  while  salts  simul- 
taneously present  acted  against  the  acids.  Acids  and  salts  were 
active  in  a  series  which  was  analogous  to  that  for  the  swelhng  and 
shrinking  of  dead  colloids. 

That  bases  caused  no  acceleration  of  sprouting  seems  to  mihtate 
against  the  original  assumption.  Borowikow  explains  this  by  the 
fact  that  the  cell  juice  in  the  growth  zone  is  essentially  acid  and  con- 
stantly forms  carbonic  acid;  the  bases  neutralize  the  acid,  forming 
neutral  unhydrated  albumin  and  in  higher  concentrations  damage 
the  plants.  In  this  way  he  explains  the  stimulatmg  action  of  dilute 
solutions  of  organic  bases  (0.001  n  urea  nitrate,  0.0015  n  caffein 
sulphate,  0.0025  n  phenylene  diamine  chlorid)  which,  according  to 
Borowikow,  act  like  their  respective  acids  since  they  are  hydrolyzed 
in  solution. 

Borowikow  expects  especially  to  bring  growth  into  relationship 
with  turgor  (tissue  distention).  Unnoticed,  great  turgor  may  be 
diminished  by  the  growth  process.  According  to  Borowikow 
growth  is  ionization  of  the  plasma  protein  by  H  ions  in  the  gro^vth 
zone,  causing  the  protein  to  pass  from  the  gel  to  the  sol  condition. 


268  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Ossification  Processes. 

One  of  the  most  interesting  of  colloid-chemical  problems  is  bone 
formation.  We  shall  see  on  page  302  that  from  an  aqueous  solution 
containing  blood  salts,  calcium  carbonate  and  calcium  phosphate 
precipitate.  The  precipitation  is  hindered  by  the  presence  of  the 
blood  colloids,  though  two-thirds  of  Ca  salts,  at  least  in  the  serum  of 
higher  animals,  occur  in  the  crystalloid  state.  This  interference 
must  stop  during  the  formation  of  bone.  To  account  for  this  there 
are  several  theoretical  possibilities :  it  may  be  assumed  that  changes  in 
the  serum  colloids  are  brought  about  at  or  from  the  bone  cells,  which 
remove  their  protective  action  and  results  in  the  precipitation  of  the 
calcium  salts.  This  agrees  with  the  views  of  Wo.  Pauli  and  Samec,* 
which  we  shall  consider  more  closely.  It  was  shown  in  their  researches 
that  the  increase  in  the  solubility  of  calcium  carbonate  by  serum 
albumin  was  475  per  cent,  and  of  calcium  phosphate  90  per  cent.  We 
would  consequently  expect  to  find  a  very  much  more  extensive  pre- 
cipitation of  calcium  phosphate  than  of  calcium  carbonate  when  the 
protective  action  was  removed.  But  in  the  case  of  bones,  the  pro- 
portions are  just  the  reverse.  The  bone  ash  of  man  contains  about 
850  parts  Ca3(P04)2  and  90  parts  CaCOs  per  1000. 

But  in  the  case  of  a  cleavage  product  of  albumin.  Wo.  Pauli 
and  Samec  found  that  the  solvent  action  upon  calcium  salts  was 
the  reverse.  Witte' s  peptone,  consisting  almost  entirely  of  albumoses, 
holds  in  solution  only  the  calcium  carbonate,  whereas  the  calcium 
phosphate  exhibits  a  diminution  in  solubility.  Based  on  these 
results,  ossification  might  occur  in  the  following  way :  In  the  bone  or 
cartilage  cells,  there  occurs  a  concentration  of  colloids  in  which  a 
large  quantity  of  calcium  salts  are  piled  up.  When  these  tissue  col- 
loids are  broken  down,  a  precipitation  occurs,  the  precipitate  consist- 
ing chiefly  of  calcium  phosphate  with  smaller  amounts  of  calcium 
carbonate.  This  corresponds  with  the  histological  evidence,  by  means 
of  which  a  tissue  destruction  may  be  seen  to  accompany  ossification. 

A  further  possibility,  which  does  not  in  the  least  contradict  the 
above  explanation  but  possibly  coincides  with  it,  is  that  phosphates 
are  set  free  and  come  into  contact  with  the  carbonates  always 
present  when  the  tissues,  especially  the  cell  nuclei,  break  down.  In 
accordance  with  well-known  physico-chemical  laws,  an  increase  in  the 
concentration  of  an  ion  (in  this  case  the  phosphate  ion)  results  in  an 
increase  in  the  calcium  phosphate  molecules,  and  this  changed  albumin 
must,  accordingly,  favor  the  precipitation  of  calcium  phosphate. 

Finally  we  may  think  of  a  kind  of  specific  adsorption  by  certain 
cell  groups.     In  fact,  M.  Pfaundler  noticed  a  selective  adsorption  of 


GROWTH,   METAMORPHOSIS  AND  DEVELOPMENT         269 

calcium  when  he  placed  pieces  of  cartilage  in  chloric!  of  üme  solution. 
This  suggests  the  method  by  which  hme  is  deposited  in  damaged 
tissues  (vessel  walls  or  tubercles).  We  must  also  consider  the 
simultaneous  precipitation  of  positively  and  negatively  charged 
albumin  with  the  breaking  down  of  calcium  salts  as  we  shall  describe 
later  when  we  discuss  concrement  formation  at  greater  length  (see 
p.  271  et  seq.). 

Finally,  we  may  consider  some  kind  of  specific  adsorption  by 
definite  cell  groups.  We  might  also  consider  the  mutual  precipita- 
tion of  positively  and  negatively  charged  albumin  which  carry  salts 
down  with  them  in  the  same  way  as  is  more  fully  described  in  the 
case  of  concrement  formation  (p.  271  et  seq.). 

What  has  been  said  here  of  bones  also  holds,  of  course,  equally  well 
in  principle  for  the  shells  of  mohuscs  and  snails,  for  the  armor  of 
crustaceans,  as  well  as  for  other  ossification  phenomena.  Morphol- 
ogists  distinguish  primarily  between  calcification  and  ossification  (see 
Gebhardt)  .  In  lower  animals  (shells  of  snails  and  mussels,  carapace 
of  crabs,  spicules  of  sponges,  etc.),  the  Hme  salt  occurs  chiefly  in 
microcrystalline  form,  as  fine  granules  in  the  calcifying  tissues.  In 
contrast  with  this  calcification,  lime  forms  an  optically  completely 
homogeneous  deposit  in  bone  and  never  occurs  as  a  formed  or 
crystalhne  precipitate.  Possibly  this  essential  difference  depends  on 
the  fact  that  special  cells,  osteophytes,  take  part  in  bone  formation. 
It  is  still  impossible  for  colloid  research  as  yet;,  to  offer  even  an 
hypothesis  in  explanation  of  this  difference. 

R.  Liesegang*  (his  correctness  in  doing  so,  I  shall  not  discuss) 
criticises  an  explanation  of  ossification  which  involves  the  presence 
of  special  cells,  the  osteoblasts.  He  calls  attention  to  the  fact  that 
deposits  of  lime  occur  in  places  where  there  are  no  osteoblasts,  as  in 
the  arterial  wall,  in  arteriosclerosis,  or  in  brain  cells.  He  evidently 
concludes  that  under  some  circumstances,  even  without  a  special 
storing-  up,  it  is  possible  to  have  a  precipitation  from  blood  serum 
supersaturated  with  calcium  salts,  in  which  action  the  formation  of 
centers  or  nuclei  possibly  take  part  (similar  to  the  theory  of  H. 
Bechhold  and  Ziegler*^  for  the  deposition  of  urates). 

The  very  marked  density  and  the  poverty  of  the  bony  framework  in 
organic  substances  is  deserving  of  special  consideration.  For  this, 
the  investigations  of  R.  Liesegang*^  offer  valuable  experimental 
support.  He  showed  that  when  calcium  phosphate  membranes 
were  allowed  to  form  in  gelatin  jellies  (by  the  diffusion  of  disodium 
phosphate  and  calcium  chlorid  towards  each  other),  that  they  were 
almost  free  from  gelatin;  to  a  certain  extent  the  organic  supporting 
substance  had  been  forced  away. 


270  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

This  investigator  has  simulated  the  formation  and  growth  of  the 
long  bones.  He  filled  test  tubes  half  full  of  gelatin  which  was  made 
alkaline,  with  tricalcium  phosphate,  for  instance;  this  layer  repre- 
sented the  periphery  of  the  bone.  After  solidification  there  was 
placed  on  top  a  thin  coating  of  gelatin  containing  a  suspension  of  tri- 
calcium phosphate;  this  was  to  represent  the  bone.  Upon  this  was 
poured  some  acid  solution,  for  instance,  lactic  acid;  representing  the 
center  of  the  bone.  The  acid  diffuses  through  the  tricalcium  phos- 
phate layer,  dissolves  the  calcium,  reaching  the  lower  periphery, 
where  the  calcium  is  precipitated  again  in  layers  as  a  phosphate.  If 
a  suitable  calcium  salt  is  added  with  the  lactic  acid,  the  layer  becomes 
stronger  and  less  porous;  in  a  successful  experiment,  it  shows,  inside, 
the  characteristic  worm-eaten  appearance  of  the  long  bones,  and  out- 
side, a  smooth  firm  and  sharply  defined  structure.  As  natural  sources 
of  acids,  R.  Liesegang  mentions  the  accumulation  of  CO2,  lactic  acid 
and  glycerophosphoric  acid  derived  from  lecithin.  [Barille  found 
that  tricalcium  phosphate  was  dissolved  by  water  containing  CO2  under 
pressure,  forming  an  unstable  compound  tribasic  calcium  carbon  phos- 
phate, CasPsOs  +  4  H2CO3  =  H2O  +  PaOsCaa :  2  CO  (CO3H)  Ca.] 

In  discussing  calcification  and  ossification  in  his  Harvey  Lecture, 
1910-11,  H.  Gideon  Wells  concludes  that  "there  seems  to  be  no 
essential  differences  between  the  processes  involved  in  normal  ossifi- 
cation and  in  most  instances  of  pathological  calcification.  Any  area 
of  calcification  may  be  changed  to  true  bone  in  the  course  of  time," 
and  that  "'  calcium  deposition  seems  to  depend  rather  on  physico- 
chemical  processes  than  on  chemical  reactions."  Jerome  Alexander 
suggests  the  importance  of  the  removal  of  or  alteration  of  protective 
substances  resulting  in  the  deposition  of  calcium  salts.  Cf.  also 
Hofmeister' s  observation  on  the  difference  between  solubility  of 
calcium  phosphate  dissolved  in  serum  and  the  dissolving  of  calcium 
phosphate  by  serum.     Tr.] 

Diseases  of  the  Bone. 

Of  the  noninfectious  bone  diseases,  rickets  and  osteomalacia  attract 
our  special  attention. 

Rickets  is  characterized  by  lime-poor,  so-called  osteoid  tissue,  in- 
stead of  the  solid  calcareous  structure.  In  this  way  a  pliable  mass 
takes  the  place  of  the  rigid  framework.  This  lack  of  lime  might 
readily  be  attributed  to  a  lack  of  lime  in  the  food,  but  it  has  been 
shown  that  this  is  certainly  not  the  cause,  since  such  a  lack  of  lime 
can  be  produced  only  by  artificial  preparation  of  the  food.  In  my 
opinion  Pauli's  theory  of  bone  formation  offers  a  good  explanation 
for  rickets.     He  supposes,  as  indicated  on  page  268,  a  prehminary 


GROWTH,  METAMORPHOSIS  AND  DEVELOPMENT         271 

tissue  degeneration;  in  rickets  we  find  quite  the  reverse,  namely, 
an  over-production  of  tlie  osteoplastic  tissue,  so  that  we  lack  the 
conditions  necessary  to  the  precipitation  of  calcium  phosphate  and 
carbonate,  or,  in  other  words,  bone  formation. 

Osteomalacia,  bone  consumption,  is  in  certain  respects  the  reverse 
of  rickets.  If  in  rickets  we  find  a  deficient  precipitation  of  insol- 
uble lime  salts,  in  osteomalacia  we  have  the  eating  away  of  existing 
bone.  Osteomalacia  occurs  most  frequently  during  pregnancy,  dur- 
ing which  even  under  normal  conditions  the  teeth  may  suffer. 
Osteoporosis,  the  bone  consumption  of  the  aged,  which  is  especially 
noticeable  in  the  skull,  belongs  to  this  group.  Here,  too,  we  must 
reject  the  theory  of  the  deficient  introduction  of  lime  salts,  as  it  is 
contradicted  by  all  metabolism  researches.  We  are  much  more  in- 
clined to  accept  a  dissolving  away  of  calcium  phosphate  and  carbon- 
ate, especially  by  acids.  Since  the  oxidizing  processes  are  deficient 
and  the  circulation  functionates  less  perfectly,  an  accumulation  of 
acids  is  not  surprising.  Magnus  Levy  has  raised  the  objection  to 
this  ''acid  theory"  that  the  proportion  of  the  calcium  phosphate  to 
calcium  carbonate  is  the  same  in  osteomalacic  as  in  normal  bones. 
He  placed  normal  bones  in  lactic  acid  and  found  that  much  more 
carbonate  than  phosphate  is  dissolved  away,  and  from  this  he  con- 
cluded that  the  ''acid  theory"  was  useless.  This  objection  can- 
not be  allowed.  If  acid  diffuses  from  any  direction  into  a  mixture 
of  calcium  carbonate  and  phosphate  imbedded  in  a  jelly,  the  acid 
advances  only  to  the  extent  that  it  has  previously  dissolved  away 
all  the  carbonate  and  phosphate;  this  was  shown  experimentally 
by  R.  LiESEGANG*^  (assuming,  of  course,  that  an  acid  stronger  than 
phosphoric  acid  is  employed).  As  may  be  readily  seen,  the  result  of 
the  experiment  depends  entirely  upon  the  conditions;  at  any  rate 
the  contribution  of  Magnus  Levy  cannot  count  against  the  "acid 
theory." 

Concrements. 

In  various  pathological  processes  we  find  in  the  body  cavities  of 
animals  and  men,  structures  varying  in  size  from  that  of  a  grain  of 
sand  to  that  of  a  fist,  and  which  have  developed  without  the  help  of 
cells.  Such  precipitates  are  called  concrements.  We  find  them  as  renal 
gravel,  urinary  calculi,  gallstones,  brain  sand  (in  the  lymph  spaces  of 
the  brain),  rice  bodies  in  the  exudate  of  diseased  joints;  as  the  pearl  of 
the  pearl  oyster;  and  similar  formations  which  are  found  at  times  in 
cocoanuts,  in  view  of  their  structure,  can  be  considered  nothing  else. 

The  common  characteristic  of  all  concrements  is  that  in  addition 
to  the  special  characteristic  ingredients  (urates  and  Cholesterin)  they 


272  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

always  contain  albuminous  elements  and  they  usually  show  a  scaly 
and  radial  structure.  These  formations  have  been  studied  with 
especial  care  by  H.  Schade*  as  well  as  by  L.  Lichtwitz. *i 

In  the  following  pages,  we  shall  consider  the  origin  of  urinary  and 
biliary  calculi  from  the  standpoint  of  these  studies. 

Urinary  Calculi.  H.  Schade  mixed  ox  plasma,  which  had  been 
made  uncoagulable  by  the  addition  of  potassium  oxalate,  with  an 
emulsion  of  calcium  phosphate  and  calcium  carbonate.  When  he 
coagulated  the  mass  by  the  addition  of  CaCl2,  a  hard  cake  formed, 
which,  when  preserved  in  salt  solution  at  40°,  shrank,  and  after  eight 
weeks  had  approximately  the  hardness  of  a  fresh  urinary  calculus. 
Fibrin  was  absolutely  necessary,  yet  from  0.07  to  0.1  per  cent  in 
plasma  diluted  ten  times  sufficed  to  produce  a  coagulation,  and  the 
phenomenon  was  the  same  if  neutral  urine  was  used  for  dilution 
instead  of  physiological  salt  solution. 

By  changing  the  composition  of  the  sediment  (mineral  ingredients) 
it  is  not  difficult  to  produce  stratified  structures  resembling  urinary 
calculi.  This  similarity  is  not  a  mere  superficial  one.  Renal  calculi 
have  been  repeatedly  found  which  were  still  soft  and  plastic  like  the 
initial  stages  of  these  artificial  stones.  Whether  the  organic  layered 
framework  of  natural  urinary  calculi  (see  Plate  II,  Fig.  51)  consists 
of  fibrin  is  still  an  open  question,  though  there  is  much  in  favor 
of  this  view.  According  to  H.  Schade  the  formation  of  urinary 
calculi  is  somewhat  as  follows:  coagulum  and  urates  sediment 
simultaneously  or  in  close  succession,  shrink  and  harden.  By  the 
repetition  of  such  processes  layers  form,  the  stone  grows,  and  after 
a  while  becomes  stony  hard,  because  the  crystalloid  ingredients  grow 
into  large  crystal  aggregates  and  take  on  a  radial  structure.  The 
lesson  for  therapeutists  is,  that  not  only  must  the  formation  of 
crystalloid  sediments  be  prevented,  but  also  the  passage  of  fibrin  or 
similar  colloids  into  the  urine.  Alone,  urinary  sediments  form  a 
crumbling  mass.  Calculus  formation  is  possible  only  by  means  of 
colloidal  "mortar." 

Gallstones  (biliary  calculi):  Gallstones  differ  very  widely  in  their 
chemical  composition.  We  recognize  those  which  merely  consist  of 
Cholesterin  and  others  which  contain  only  calcium  bilirubin;  between 
these  extreme  forms  occur  all  sorts,  consisting  of  mixtures  of  these 
two  chief  constituents  with  albuminous  material. 

Without  going  into  the  individual  reasons,  it  may  be  said  that, 
according  to  H.  Schade,  Cholesterin  appears  to  be  dissolved  in  the 
bile  as  a  hydrophile,  and  calcium  bihrubin  as  a  hydrophobe  colloid. 
It  must  also  be  noted  that,  besides  the  chelates,  the  bile  contains 
salts  of  the  fatty  acids,  lecithin  and  mucinous  substances  which 


GROWTH,  METAMORPHOSIS  AND  DEVELOPMENT        273 

must  partly  be  regarded  as  solvents  for  the  gallstone  material. 
Cholesterin  precipitates  in  individual  crystals  from  a  supersaturated 
aqueous  solution  of  etiolates;  however  a  few  drops  of  an  oil  suf- 
fice to  cause  precipitation  in  an  amorphous  clump  very  similar  to 
the  ''myelin  clump"  which  Naunyn  described  as  the  (uranlage) 
'' precursor  of  gallstones"  (Plate  II,  Fig.  52).  After  a  few  days, 
radiating  crystallization  starts  from  the  center  (Plate  II,  Fig.  53), 
oil  droplets  are  released  and  may  again  give  rise  to  oil-cholesterin 
precipitates.  In  this  way  may  be  prepared,  artificially,  hard  or  more 
or  less  plastic  Cholesterin  stones,  such  as  rarely  occur,  however,  in  the 
gall  bladder.  The  presence  of  fats  or  fatty  acids  is,  therefore,  a 
requisite  for  their  formation.  For  the  precipitation  of  Cholesterin 
it  is  only  necessary  that  the  substances  which  hold  it  in  solution, 
the  cholates  and  salts  of  the  fatty  acids,  be  destroyed.  A  priori, 
a  supersaturation  with  Cholesterin  is  brought  about  in  all  processes 
that  interfere  with  the  normal  alkaline  reaction  of  the  bile,  especially 
infection  with  B.  coli,  B.  typhosus,  B.  pyocyaneus  and  B.  proteus. 
I  am  inclined  to  accept  the  view  of  L.  Lichtwitz*i  that  the  acid  for- 
mation of  these  bacteria  is  chiefly  responsible  for  the  breaking  down 
of  cholates  and  soaps,  since  staphylococcus  aureus,  which  does  not  form 
acid,  causes  no  separation  of  Cholesterin.  The  precipitation  of 
Cholesterin  can  also  be  caused  by  sterile  autolysis  as  well  as  by 
rendering  less  favorable  the  conditions  requisite  to  solution,  since 
cholates  are  absorbed  by  the  walls  of  the  gall  bladder  if  the  bile 
stagnates  there  (congestion). 

Bilirubin  forms  amorphous  precipitates  with  lime  salts  which 
normally  do  not  sediment  out  in  the  bile.  In  the  presence  of  al- 
bumin and  fibrin,  under  conditions  as  yet  not  accurately  studied, 
calcium  bilirubin  may  precipitate  and  include  the  albuminous  in- 
gredients, giving  rise  to  clumps  which  in  their  cheesy  stt-uctures 
are  very  like  natural  calcium  bilirubin  stones  (Plate  II,  Fig.  54). 
In  my  opinion  the  neutral  or  faintly  alkafine  reaction  of  the  bile 
is  essential  for  the  development  of  a  calcium  bilirubin  stone  as  op- 
posed to  the  Cholesterin  stones,  which  require  acidity.  H.  Schade* 
considers  that  catarrhs,  inflammatory  and  strongly  exudative  proc- 
esses in  which  much  lime  enters  the  bile  are  responsible  for  the 
formation  of  calcium  bilirubin  stones;  this  view  explains  the  occur- 
rence in  them  of  albuminous  ingredients. 

In  some  of  the  so-called  mixed  forms  there  may  be  an  alternation 
of  processes  which  condition  the  separation  of  Cholesterin  and  of 
calcium  bilirubin. 

An  answer  to  the  question  whether  a  simultaneous  precipitation  of 
Cholesterin  and  calcium  bilirubin  may  occur  would  be  very  interesting. 


274  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

H.  Schade  rightly  sees  in  every  section  through  a  stone,  a  ''valu- 
able document  written  by  nature  on  the  development  and  course 
of  gallstone  disease. " 

Gout. 

Gouty  deposits,  tophi,  which  form  in  the  joints  by  preference, 
offer  a  certain  analogy  to  ossification  processes.  In  uric  acid  arthri- 
tis as  well,  the  blood  is  from  time  to  time  supersaturated  with  a 
difficultly  soluble  salt,  monosodium  urate,  which  precipitates  under 
favorable  conditions. 

Gout  is  regarded  as  a  disease  of  nuclein  metaboHsm.  We  must 
forego  a  discussion  of  this  part  of  the  question  at  present,  and  limit 
ourselves  to  determining  the  influence  exerted  by  the  colloids  of  the 
organism  upon  the  deposition  of  urates,  the  most  characteristic 
phenomenon  in  the  disease  picture  of  uric  acid  arthritis.  Two 
papers  by  H.  Bechhold  and  J.  Ziegler*''  as  well  as  by  F.  Gudzent,* 
which  appeared  in  1912,  have  given  a  certain  direction  to  these 
views. 

Until  the  appearance  of  these  papers,  no  stress  was  laid  on  the 
question  whether  uric  acid  appeared  as  such  or  as  urates  in  the  organ- 
ism. The  reason  for  this  was  that  all  analytical  investigations  deter- 
mine the  uric  acid,  and  since  it  was  known  that  uric  acid  is  much  less 
soluble  in  water  than  its  alkaline  salt,  it  was  taken  for  granted  that 
the  same  condition  held  for  the  fluids  of  the  body,  especially  for  the 
serum.  H.  Bechhold  and  J.  Ziegler  showed,  however,  that  no 
free  uric  acid  but  only  urates,  chiefly  sodium  urate,  exist  in  the  body, 
and  they  showed,  moreover,  that  sodium  urate  was  much  less  solu- 
ble in  serum  than  is  uric  acid.  The  subsequent  publication  of 
F.  Gudzent  confirmed  and  explained  this  experimental  result  by  the 
electrolytic  dissociation  of  the  electrolytes  in  question.  F.  Gudzent 
started  with  the  idea  that  albuminous  substances  play  no  part  in 
these  processes,  but  H.  Bechhold  and  J.  Ziegler  have  shown  that 
this  assumption  is  erroneous.  A  few  figures  will  explain  what  has 
been  stated. 


Solubility  (at  37°  C). 

In  water. 

In  serum. 

1  :  15500 
1  :      665 

Uric  acid                  

J  1 :     1925  (a) 
1  1  :  40,000  (6) 

Monosodium  urate^         

1  In  dissolving  sodium  urate  in  these  two  ways,  we  do  not  obtain  the  same  equilibrium.  Only 
1  :  40,000  monosodium  urate  dissolves  in  serum  (&),  but  if  monosodium  urate  is  permitted  to  form  by 
dissolving  uric  acid  in  serum  (a),  then  1  :  1925  dissolves.  [This  is  probably  due  to  the  protective  ac- 
tion of  the  serum,  as  the  result  of  which  some  of  the  sodium  urate  remains  colloidally  dispersed.    Tr.] 


GROWTH,   METAMORPHOSIS  AND  DEVELOPMENT         275 

It  follows  from  this  in  contradiction  to  previous  views  that  the 
blood  in  gout  is  frequently  supersaturated  with  monosodium  urate. 
According  to  the  analysis  of  G.  Klemperer,  Magnus-Levy  and 
SalOxMon,  the  uric  acid  content  of  the  blood  in  gout  varies  between 
30  and  80  mg.  per  liter,  whereas  in  normal  blood  at  most  only 
traces  of  uric  acid  can  be  demonstrated.  When  the  content  reaches 
25  mg.  of  sodium  urate  per  liter  of  blood  serum,  every  further  addi- 
tion must  be  associated  with  a  deposition  of  sodiimi  urate,  provided 
urate  nuclei  are  present.  We  thus  see  that  the  serum  colloids  are 
of  great  importance  for  the  solution  of  sodium  urate  in  the  blood 
and  in  preventing  its  deposition  in  gouty  processes.  [Stanley  R. 
Benedict  in  his  Harvey  Lecture,  1915-1916,  p.  362,  discusses  the 
presence  of  two  forms  of  uric  acid  in  blood.  He  determined  ten 
times  the  amount  of  uric  acid  originally  obtained  by  the  preliminary 
boiling  of  the  protein  free  filtrate  with  hydrochloric  acid.  The  prob- 
able destruction  of  a  ''  protective "  substance  is  quite  apparent. 
This  aspect  has  an  important  bearing  on  uric  acid  determinations  in 
nephritis  and  out.     Tr.] 

The  influence  on  these  processes  exerted  by  radium  emanations 
which  inhibit  the  deposition  of  sodium  urate  from  supersaturated 
serum  (H.  Bechhold  and  J.  Ziegler**)  deserves  the  attention  of 
students  of  colloids. 


CHAPTER  XVI. 

THE   CELL. 

It  is  said  that  the  cells  are  the  structural  units  of  the  body;  this 
comparison,  however,  is  valid  only  to  a  very  limited  extent.  Build- 
ing stones  do  not  vary  much  in  form  and  structure,  and  especially 
in  their  individual  uses;  the  cells  of  the  body,  however,  are  of  such 
manifold  appearance  and  have  such  numerous  uses  that  it  is  very 
difficult  to  discover  what  is  common  to  the  various  kinds.  A  cell 
may  consist  of  protoplasm,  the  cell  contents  and  a  nucleus;  there 
may  even  be  no  nucleus. 

In  plants,  there  is  usually  a  visible  cell  pellicle  which  supports  the 
protoplasm;  this  occurs  less  frequently  in  animal  cells.  Protoplasm, 
nucleus,  and  pellicle  are,  however,  the  microscopically  distinguisha- 
ble parts.  Theory  requires  an  invisible  plasma  pellicle  which  is  con- 
sidered the  surface  of  the  protoplasm. 

There  can  be  no  doubt  that  a  cell  consists  of  many  elements 
beneath  the  limits  of  visibility,  which  are  responsible  for  definite 
functions.  Independent  microorganisms  exist  which  are  invisible 
because  of  their  size  (the  ultramicroscopic  causes  of  some  diseases, 
e.g.,  smallpox,  measles,  etc.),  which  possess  all  the  properties  of  an 
independent  complicated  organism  (nutrition,  propagation,  etc.). 
We  shall  endeavor  to  get  an  idea  of  the  molecular  structure  of  a 
cell  from  an  analogy  of  F.  Hofmeister  who  took  as  his  example  a 
liver  cell,  which  performs  particularly  numerous  functions.  It  occu- 
pies approximately  the  space  of  a  cube  with  edges  20  /x  long  =  8000  /J 
=  8'10~^  mm^.  Assuming  the  following  conditions  F.  Hofmeister 
arrives  at  the  subsequent  figures.  A  gram  molecule  of  any  chemical 
substance  consists  of  0.62  quadrillion  (0.62«  10^*)  molecules.  From 
this  0.62- 10^*  we  may  calculate  the  number  of  molecules  present  in 
a  definite  space  if  we  know  the  weight  and  composition  of  a  cell. 

F.  Hofmeister  assumes  that  for  colloids,  on  the  average,  the 
molecular  weight  is  that  of  hemoglobin  (about  16,000),  that  for 
lipoids  800,  and  for  crystalloids  100. 

Consequently,  we  must  figure  that  a  liver  cell  contains 

76  per  cent  water 225,000  milliard  molecules^ 

16  per  cent  protein 53        "  " 

2^  per  cent  lipoids 166        "  " 

5|  per  cent  crystalloids 2,900        "  " 

1  [1  milliard  =  1000  millions.      Tr.] 
276 


THE  CELL  277 

In  order  to  grasp  these  figures  F.  Hofmeister  shows  how  a  struc- 
ture might  be  erected  whose  molecules  are  bricks,  not  to  exceed  in 
number  200,000  miUiards,  of  which  200  milliards  colloid  molecules 
with  a  portion  of  the  salt  molecules  form  the  walls,  roof,  ceihngs, 
etc.,  whereas  the  water  molecules  with  the  remaining  crystalloid 
molecules  fill  the  rooms,  halls  and  corridors.  If  such  a  structure  had 
the  enormous  average  height  of  50  meters  it  would  cover  a  ground 
space  7000  square  kilometers  or  one-half  the  area  of  Alsace-Lorraine. 
It  is  evident  that  the  compexity  of  the  molecular  structure  of  a  cell 
baffles  our  powers  of  description. 

A  cube  with  edges  O.l^u  which  is  much  smaller  than  the  Hmits  of 
microscopic  visibility  contains  25  million  molecules  of  water,  25 
thousand  molecules  of  colloidal,  and  250  thousand  molecules  of 
crystalloidal  substance,  which  under  the  same  conditions  would  cor- 
respond to  a  building  100  meters  front,  20  meters  high  and  20  meters 
deep. 

Protoplasm. 

Until  recently  there  was  little  definite  knowledge  concerning  the 
colloidal  nature  of  protoplasm,  that  is,  whether  it  was  fluid  or  gela- 
tinized. It  was  known  that  after  the  fragmentation  of  yeast  cells  it 
was  possible  to  press  out  a  juice  containing  various  enzymes,  and 
that  meat  juice  obtained  in  a  similar  manner  contained  albumin. 
In  the  case  of  yeast  it  may  be  inferred  that  the  protoplasm  contains 
sols,  but  in  the  case  of  muscle  such  an  inference  is  met  by  the  objec- 
tion that  the  albuminous  substance  may  have  arisen  from  the  blood 
serum  which  bathes  the  muscle  fibers.  The  facts  that  portions  of 
cells  form  drops  and  that  foreign  fluids  in  protoplasm  assume  spherical 
shapes  likewise  point  to  the  fluid  nature  of  many  protoplasms. 

Most  of  the  numerous  investigations  concerning  the  physical 
nature  of  protoplasm  are  at  present  of  mere  historical  interest, 
since  the  ultramicroscope  has  solved  many  of  the  main  questions  or 
has  placed  us  in  a  position  to  do  so  in  the  future.  One  of  the  most 
important  criteria  for  differentiating  between  a  sol  and  a  gel  ^  is  the 
presence  of  Brownian  movement.  If  it  is  possible  to  observe  an 
oscillatory  movement  in  the  granules  of  a  cell,  such  granules  must 
be  in  a  fluid  medium;  if  they  are  motionless  the  medium  must  be 
either  a  gel  or  very  viscous.  If  we  observe  that  the  oscillating 
movement  has  ceased,  it  means  that  the  fluid  has  gelatinized. 

Numerous  ultramicroscopic  observations  of  cells  have  been  pub- 
lished. Plant  cells  have  been  studied  most  carefully  by  N.  Gaid- 
UKOV.*  He  studied  the  pollen  hairs  of  tradescantia,  m3':xom5^cetes 
(shme  fungi),  the  ceils  of  various  algae  (spirogyra,  cladophora,  oedogo- 

1  [There  is  no  sharp  Une  between  sol  and  gel.  The  more  viscous  the  medium 
the  longer  time  the  changes  take,  vid-  metals  for  examples.     Tr.] 


278  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

nium,  desmids,  diatoms,  oscillaria,  etc.),  yeasts,  the  common  frog- 
bit,  vallisneria,  and  several  mosses. 

Gaidukov  comes  to  the  following  conclusion  as  the  result  of 
these  investigations:  Protoplasm  must  consist  of  hydrosols  because 
everywhere  in  living  protoplasm  he  saw  particles  with  Brownian 
movement.  Frequently  he  observed  that  the  particles  combined 
or  separated  and  that  their  number  increased  or  diminished,  phe- 
nomena which  in  my  opinion  represented  metabolic  changes.  In 
some,  usually  in  very  well  nourished  cells,  there  were  no  movements, 
which  may  be  attributed  to  the  fact  that  the  distances  between  the 
numerous  particles  were  too  small. 

A  transition  from  sol  to  gel  condition,  i.e.,  the  cessation  of  Brownian 
movement,  was  not  observed  in  normal  living  cells.  The  colloids  of 
plant  protoplasm  evidently  consist  of  a  reversible  and  an  irreversible 
portion.  If  a  cell  is  injured  so  that  protoplasm  escapes,  a  portion 
will  expand  in  the  water  and  ultimately  be  dissolved  in  it,  whereas 
other  portions  combine  (precipitate).  This  was  observed  in  living 
and  in  dead  protoplasm.  The  observation  of  0.  Nägeli  is  analogous: 
If  we  crush  a  root  hair  of  hydrocharis  in  water  under  a  cover  glass, 
clumps  of  protoplasm  pass  through  the  rent.  They  are  immediately 
surrounded  by  a  membrane  which  is  impermeable  for  dyes;  i.e.,  they 
are  coagulated  on  the  surface.  There  is  formed  at  the  site  of  the 
wound  an  irreversible  layer  of  hydrogel  similar  to  the  fibrin  forma- 
tion of  higher  animals.  The  ultramiscoscopic  observations  quoted 
entirely  confirm  what  was  earlier  observed  when  plant  cells  absorbed 
water.  For  instance,  if  we  place  myxomycetes  in  water,  they  swell; 
and  in  spite  of  the  increase  in  surface,  the  outer  hyaloplasm  retains 
its  thickness.  Evidently  the  entering  water  causes  a  gelatinization 
of  the  granular  plasma  at  the  surface  of  contact,  so  that  the  hyalo- 
plasm layer  spontaneously  supplements  itself. 

W.  W.  Lepeschkin  regards  protoplasm  as  a  loose  combination  of 
proteins  and  lipoids  which  breaks  down  under  lethal  conditions 
(coagulation).  If  I  understand  him  correctly  he  does  not  assume 
the  existence  of  a  plasma  pellicle  (see  p.  245)  but  believes  that  all 
the  properties  ascribed  by  other  investigators  to  this  membrane  as 
a  limiting  surface  should  be  attributed  to  the  protoplasm  as  a  whole. 

It  must  be  remembered  that  all  these  observations  experience 
certain  limitations  depending  on  the  kind  and  the  part  of  the  plant 
involved.  As  a  result  of  his  experiments  with  sunflower  seedlings, 
BoRowiKow  assumes  that  plasma  exists  in  seeds  and  spores  as  a 
solid  phase  which  changes  to  the  gel  condition  in  resting  plants 
(evidently  jellies  are  meant). 

In  the  growth  period,  the  plasma  as  the  result  of  hydration  exists 


THE  CELL  279 

in  the  gel  condition,  the  one  in  which  we  usually  find  it  in  cells 
according  to  Borowikow. 

With  the  occurrence  of  death  -protoplasm  gelatinizes,  Brownian  move- 
ment of  the  smaller  particles  ceases,  and  the  structure  of  the  gel 
appears  in  the  ultramicroscope  as  a  conglomeration  of  many  re- 
flecting platelets.  It  makes  a  substantial  difference  whether  the 
protoplasm  slowly  dies  or  is  suddenly  killed  by  a  fixative  (alcohol, 
formalin,  etc.)-  In  the  first  instance  there  is  a  precipitation  (floccu- 
lation),  whereas,  in  the  latter  there  is  a  stiffening;  this  difference 
may  be  readily  recognized  under  the  ultramicroscope. 

From  this  we  may  understand  why  a  dead  plant  cell  simply  bursts 
in  water,  for  the  defects  are  no  longer  repaired  from  within.  The 
cell  contents  have  been  already  gelatinized.  Chlor ohlasts  (chlorophyl 
granules)  may  be  assumed  to  possess  colloidal  properties  similar  to 
protoplasm ;  only  it  seems  the  latter  are  more  delicate  (Ponomarew)  . 
The  living  protoplasm  of  many  animal  cells,  however,  seems  to  exist 
as  a  gel.  At  least  in  monocellular  organisms,  blood  cells,  etc.,  A. 
Mayer  and  G.  Schaeffer*  could  not  discover  any  Brownian  move- 
ment of  certain  granules. 

On  account  of  the  great  differentiation  of  animal  cells,  more  com- 
prehensive investigations  must  be  awaited;  thus  it  appears  to  me 
probable  that  red  blood  cells  have  viscous  contents  (see  p.  305). 

The  Nucleus. 

We  know  even  less  about  the  colloidal  nature  of  the  nucleus  than 
of  protoplasm.  Ultramicroscopically,  the  nucleus  appears  to  be  a 
complex  of  hydrosols  containing  larger  particles  and  to  be  quite 
poor  in  water.  This  corresponds  well  with  the  picture  produced  by 
staining. 

The  colloids  of  cell  protoplasm  seem  to  be  rather  indifferent 
chemically;  they  are  poorly  stained  by  both  acid  and  basic  dyes. 
The  nucleus,  or  more  properly  the  chromatin  substance,  seems  to 
possess  pronounced  acid  properties,  which  are  manifested  by  its 
intense  staining  with  basic  dyes  (see  A.  Kossel*). 

The  Cell  Membrane  and  the  Plasma  Pellicle. 

The  cell  pellicle  imparts  its  shape  to  the  fluid  protoplasm  which 
otherwise  would  be  spherical  as  the  result  of  surface  tension.  The 
cell  pellicle  occurs  in  plants  especially.  In  animals  an  interior 
skeleton  or  a  spongy  framework  may  determine  shape.  Theory 
requires  an   additional   invisible   plasma   pellicle   as  bounding   the 


280  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

protoplasm.  Formerly  from  an  interpretation  of  the  experimental 
facts  as  pure  osmosis,  the  pellicle  was  considered  semipermeable, 
that  is,  permeable  for  water  but  impermeable  for  everything  else. 
This  view  is  theoretically  untenable  since  the  cell  requires  numerous 
substances  and  there  must  also  be  an  exit  for  excreta.  As  a  matter 
of  fact,  the  studies  of  recent  years  have  shown  that  the  plasma 
pellicle  is  by  no  means  as  impermeable  for  many  substances  as  was 
assumed.  The  attempt  has  been  made  to  decide  its  chemical  and 
physical  condition  from  the  nature  of  the  substances  which  pass 
through  it. 

The  statements  on  page  281  show  that  there  is  no  uniform  opinion 
as  to  whether  the  plasma  pellicle  consists  of  albuminous  or  lipoid 
substance  or  a  mixture  of  both.  In  my  opinion  it  differs  in  each 
instance  depending  on  the  contents  of  the  cell.  However,  colloid 
research  offers  at  least  a  foundation  for  a  conception  of  the  plasma 
pellicle. 

In  animal  cells,  with  few  exceptions,  we  can  discover  no  membrane, 
yet  many  of  their  properties  indicate  that  they  also  possess  some  sort 
of  a  pellicle.  Colloid  chemistry  gives  us  a  basis  for  the  explanation  of 
such  phenomena.  The  conditions  are  most  simple  when  the  cehs  are 
surrounded  by  air.  We  know  from  page  33  that  colloids  concentrate 
and  unite  into  a  firm  skin  at  an  interface,  fluid/air.  The  process  is 
much  more  complicated  in  the  case  of  cells  in  a  fluid  or  semifluid 
medium.  Let  us  recah  the  following  experiment:  If  ether  is  shaken 
with  water  containing  albumin  or  albumose,  there  will  form  a  foam 
consisting  of  drops  of  ether  surrounded  by  albumin  or  albumose  films. 
Certain  other  colloids  and  fluids  permit  the  formation  of  fluid  foams, 
which  have  unmistakable  similarities  to  agglomerations  of  cells. 

Although  this  analogy  may  at  first  sight  seem  entirely  superficial, 
we  must  remember  that  the  interface  between  two  immiscible 
fluids  and  between  a  fluid  containing  solid  or  semisolid  bodies  (gel), 
possesses  other  properties  than  does  the  interior  (see  pp.  14-17, 
et  seq.).  The  surface  of  a  cell  must  have  special  properties;  such  sub- 
stances as  shall  lower  the  surface  tension  must  collect  there.  These 
substances  are  probably  lipoids  (lecithin  and  Cholesterin)  which  have 
been  demonstrated  in  every  cell,  animal  as  well  as  vegetable.  The 
thickness  of  the  transition  layer  varies,  according  to  different  ob- 
servers of  different  substances,  from  1  to  25  /jl/jl;  whereas  the  thick- 
ness of  the  material  coherent  pellicle  (lecithin,  etc.)  does  not  need  to 
be  thicker  than  from  0.3  to  7  fxfjL.  These  are  minimal  figures  which 
show  that  a  membrane  may  be  entirely  invisible  with  the  microscope 
and  yet  fulfil  all  the  conditions  of  a  true  membrane  as  far  as  the 
transfer  of  material  is  concerned. 


.       THE  CELL  281 

In  brief  our  conclusions  so  far  are:  Every  cell  at  its  surface 
possesses  a  membrane  which  is  dependent  upon  the  composition  of 
the  interior  of  the  cell.  This  membrane  may  be  visible  and  may 
have  been  formed  through  the  gelatinization  of  the  cell  protoplasm  at 
the  periphery.  It  may,  on  the  other  hand,  be  so  thin  as  to  be  in- 
visible, being  formed  by  the  concentration  and  spreading  out  of  such 
albuminous  and  fatty  colloids  as  diminish  the  surface  tension  of  the 
cell  content  at  the  interface.  The  cell  membranes,  developing  as  a  result 
of  the  gelatinization  of  cell  protoplasm,  are  at  first,  in  youth,  expansile 
and  elastic;  with  increasing  age  these  membrane  colloids,  depending 
upon  their  environment  and  upon  chemical  influences,  or  as  a  result 
of  mere  colloid  aging  phenomena,  become  poor  in  water  and  lose  their 
elasticity. 

[In  his  "  Growth  and  Form,"  Cambridge,  1917,  D'Arcy  W.  Thomp- 
son invokes  the  aid  of  colloid  phenomena  in  discussing  the  dynamics 
of  cell  Hfe.     Tr.j 


CHAPTER  XVII. 

THE   MOVEMENTS   OF   ORGANISMS. 
The  Movements  of  Lower  Organisms. 

Freedom  of  the  will  is  still  a  problem  in  philosophy,  and  even  the 
investigation  of  the  purely  reflex  phenomena  and  actions  of  higher 
organisms  is  still  entirely  in  the  stage  of  observation  and  measure- 
ment. In  any  case,  it  is  still  impossible  to  connect  the  external 
stimulus  and  the  resultant  action  by  a  series  of  obvious  physical  and 
chemical  processes. 

It  is  otherwise  in  the  case  of  the  movements  of  certain  portions  of 
plants  and  of  the  lowest  organisms,  especially  certain  amebse  and 
their  relatives,  our  symbiotic  blood  fellows,  the  leucocytes.  In  this 
case,  opportunities  are  offered  for  an  exact  explanation  of  their 
movements  and  actions;  but  even  here  analogies  must  frequently 
carry  us  over  gaps. 

It  is  customary  to  refer  to  such  regulated  movements  of  lower 
organisms  and  portions  of  plants  as  tropisms.  [In  this  connection 
reference  should  be  made  to  the  discussion  on  "Animal  Instincts  and 
Tropisms  in  the  Organism  as  a  Whole,"  by  Jacques  Loeb.  G.  P. 
Putnam's  Sons,  1916.  John  Hays  Hammond,  Jr.,  has  constructed 
heliotropic  machines  which  follow  a  lantern  in  the  dark.  The 
"  retina  "  consists  of  selenium  wire  which  changes  its  galvanic  resist- 
ance when  illuminated.  Tr.]  We  speak  of  heliotropism  when  certain 
plankton  organisms  swim  toward  the  light  or  when  a  tree  or  a  flower 
grows  toward  the  light.  We  speak  of  positive  thermotropism  if  a  root 
grows  in  the  direction  of  a  heat  stimulus,  of  negative  thermotropism 
when  it  grows  away  from  it.  Every  fact  in  this  connection  is  not 
only  valuable  in  explaining  the  subject  but  serves  as  well  to  enrich 
the  meaning  of  the  term  ''stimulus."  "Stimulus"  is  an  expression 
employed  in  biology  wherever  the  more  profound  causes  are  not 
evident. 

Martin  H.  Fischer  has  already  indicated  how  tropisms  may  be 
explained  in  analogy  to  curling  sheets  of  gelatin. 

Th.  Parodko  contributed  extremely  valuable  studies  on  plant 
tropisms.  He  stimulated  growing  roots  from  one  side  and  they 
became  crooked.  The  stimuli  were  chemicals,  heat  and  traumata. 
He  concluded  that  all  these  tropisms  might  be  explained  by  protein 

282 


THE  MOVEMENTS  OF  ORGANISMS  283 

coagulation  in  the  affected  cells.  All  substances  and  concentrations 
which  salt  out  or  precipitate  protein  proved  to  be  chemotropic.  In 
connection  with  positive  and  negative  chemotropism,  salts  of  the 
alkalis  and  earth  alkalis  could  be  arranged  in  a  lyotropic  series  similar 
to  that  we  have  repeatedly  found  in  the  precipitation  of  albumin  and 
the  swelling  of  gelatin,  fibrin,  etc.  The  salts  of  the  heavy  metals 
act  still  more  strongly  and  always  negatively  chemotropic. 

Those  movements  which  are  manifest  as  general  effects  are  still  the 
most  accessible  to  investigation. 

When  placed  between  electrodes,  bacteria,  spermatozoa,  yeast  cells 
and  red  and  white  blood  corpuscles  migrate  to  the  anode;  amebse 
pass  to  the  cathode.  Although  organized  suspensions  and  colloids 
migrate  either  to  the  anode  or  a  few  {e.g.,  iron  oxid  hydrosol  and 
aluminium  oxid  hydrosol)  to  the  cathode,  hydrophile  organic  colloids 
such  as  thoroughly  dialyzed  albumin  and  gelatin  pass  in  no  definite 
direction  when  placed  in  an  electric  field;  they  acquire  a  definite 
direction  only  by  the  addition  of  electrolytes.  OH  ions  cause  an 
anodal  and  H  ions  a  cathodal  migration.  Since  the  organisms 
mentioned,  considered  as  a  whole,  are  hydrophile  organic  colloids, 
we  must  assume  that  their  direction  of  migration  in  the  electric  field 
is  determined  by  the  ions  clinging  to  them.  Normal  albumin  with 
a  content  up  to  0.01  normal  NaHCOs  still  migrates  to  the  anode. 
We  need  not  be  surprised,  therefore,  that  the  majority  of  micro- 
organs  and  microorganisms  also  migrate  to  the  anode.  The  problem 
reduces  itself  to  determining  the  direction  taken  by  pure  albumin. 

The  cathodal  migration  of  amebse  is  remarkable  and  requires  more 
thorough  study.  Equally  remarkable  is  the  fact  observed  by  H. 
Bechhold,*  as  well  as  by  M.  Neisser  and  U.  Friedemann,*  that 
agglutinated  bacteria  lose  their  direction  of  migration  (p.  205), 
agglutinin  having  produced  a  neutralization. 

Following  the  ideas  of  G.  Berthold,*  we  are  nowadays  tempted  to 
explain  by  a  simple  formula  certain  individual  movements  of  the 
lower  organisms  and  of  leucocytes;  that  is,  by  changes  in  surface 
tension.^  A  fluid  or  semifluid  structure  which  is  constantly  under  the 
stress  of  surface  tension  assumes  a  spherical  form,  as,  for  instance,  oil 
in  a  mixture  of  alcohol  and  water.  If  such  a  drop  is  placed  between 
two  other  phases,  a  change  in  form  occurs,  and  mth  it  a  movement. 
A  drop  of  oil  on  the  surface  of  water  spreads  out;  every  moistening 
brings  about  an  enlargement  of  the  surface,  a  spreading  out  upon  the 
moistened  body  (see  p.  17).  A  structure  may  suffer  a  change  of 
surface  tension  in  some  single  spot,  locally,  so  that  a  movement 

^  A  full  bibliography  is  given  in  L.  Rhumbler's  "  Zur  Theorie  der  Oberflächen 
kräfte  der  Amöben":  Zeitschr.  f.  wissensch.  Zoologie,  83. 


284  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

occurs;  this  may  be  induced  for  instance  by  an  electric  charge, 
chemical  reactions  and  the  like.  Thus  a  structure  may  retain  its 
general  spherical  form  yet  increase  its  surface  at  some  single  point, 
flattening  out,  putting  out  limbs,  pulsating,  making  slight  movements 
which  may  be  explained  purely  by  physical  chemistry.  The  vital 
phenomena  of  amebse  and  of  leucocytes  which  are  evidenced  espe- 
cially by  movements  of  the  plasma  may  be  regarded  as  changes 
in  surface  tension.  Portions  of  plasma  (pseudopodia)  are  far  ex- 
tended and  the  remainder  of  the  body  follows  them,  so  that  move- 
ments of  progression  arise.  Sometimes  the  pseudopodia  surround 
foreign  bodies,  a  starch  granule,  a  bacterium  or  the  hke  and  draw  it 
into  the  ameba  or  leucocyte;   ingestion  of  food  thus  takes  place. 

The  migrations  of  an  ameba,  according  to  L.  Rhumbler,  may  be 
deceptively  imitated  with  a  drop  of  chloroform  in  the  following 
way:  A  Petri  dish  is  covered  with  an  alcoholic  solution  of  shellac 
and  the  excess  is  poured  off,  so  that  after  a  few  minutes  the  shellac 
layer  is  superficially  hardened.  Boiled  water  is  then  poured  into  the 
dish  and  a  drop  of  chloroform  dropped  on  the  shellac  with  a  pipette. 
Immediately  the  drop  begins  its  characteristic  migration,  especially 
if  it  is  pushed  with  a  glass  rod  inserted  between  the  chloroform  and 
the  shellac  layer.  The  phenomenon  is  explained  as  follows :  a  marked 
surface  tension  develops  between  the  chloroform,  the  water  and  the 
moist  shellac  layer;  soon  chloroform  and  shellac  commence  to  be 
moistened  at  some  point  and  at  this  point  the  surface  tension  of 
the  chloroform  is  lowered  and  it  seeks  to  spread  itself  out.  In  this 
way  the  chloroform  drop  progresses  in  a  way  similar  to  the  flatten- 
ing of  the  advancing  margin  of  an  ameba.  The  thin  shellac  layer 
is  dissolved  by  the  chloroform  flowing  over  it,  so  that  the  path 
traversed  by  the  drop  ''appears  as  if  cut  out  of  the  shellac."  Still 
more  deceptive  is  the  similarity  of  movement  if  one  does  not  take  a 
surface  entirely  covered  with  shellac,  but  prescribes  the  path  of  the 
drop  by  a  fine  shellac  line  and  retards  the  movements  by  the  ad- 
dition of  Canada  balsam  or  neat's-foot  oil  to  the  chloroform.  Ac- 
cording to  the  proportions  of  chloroform,  size  of  drop,  thickness  of 
the  shellac  layer  and  the  degree  of  its  dryness,  the  movements  may 
imitate  the  most  diverse  kinds  of  amebse.  If  a  drop  of  chloroform 
is  placed  on  a  spot  of  shellac  which  branches  in  various  directions, 
an  imitation  of  the  spreading  of  pseudopodia  is  obtained.  The  tak- 
ing up  of  nourishment  (taking  up  of  oscillaria  threads  by  ameba 
verrucosa)  may,  according  to  L.  Rhumbler,  be  imitated  when  a  drop 
of  chloroform  in  water  comes  into  contact  with  a  thread  of  shellac; 
the  drop  completely  envelops  the  thread  of  shellac  and  rolls  it  up 
into  itself. 


THE  MOVEMENTS  OF  ORGANISMS  285 

At  times  small  amebse  are  pursued  by  larger  ones,  the  former 
change  their  direction  and  their  speed,  the  pursuer  continues  its 
journey  and  catches  its  prey,  which  may  again  escape,  and  the 
pursuit  continues.  All  these  processes  are  explained  according  to 
L.  Rhumbler,  without  invoking  a  conscious  intelligence  and  pur- 
poseful movements,  by  the  trail  left  beliind  by  the  pursued  ameba,  just 
as  the  chloroform  drop  pursues  the  track  of  shellac  mentioned  above. 

Though  the  movements  are  so  similar  and  the  explanation  by 
changing  surface  tension  is  so  clear,  we  are  still  forced  to  enquire 
how  the  surface  tension  of  amebse  and  leucocytes  is  changed.  An- 
alogy is  quite  absent  in  the  character  of  the  substances  whose  sur- 
faces are  in  contact  and  in  the  physical  process  (solution  of  the  shellac) 
that  takes  place.  It  was  assumed  that  substances  which  diminish 
surface  tension  (for  instance,  soaps,  albuminates,  L.  Michaelis)  form 
at  the  point  of  motion  and  then  break  up  again.  Though  a  definite 
demonstration  has  not  been  possible,  I  shall  discuss  an  hypothesis  of 
L.  Hirschfeld*  which  has  much  to  recommend  it  in  certain  cases. 
We  know  that  an  electric  charge  depresses  the  surface  tension  (see 
p.  87)  but  the  question  is  whether  the  development  of  an  electrical 
charge  at  any  point  of  a  mass  of  protoplasm  is  conceivable.  Let 
us  consider  the  circumstances  under  which  a  bacterium  approaches 
an  ameba  that  puts  out  a  Pseudopodium,  envelops  the  bacterium 
and  draws  it  in.  Between  two  electrodes,  amebse  migrate  to  the 
cathode  and  bacteria  to  the  anode.  H  ions  diminish  surface  ten- 
sion, causing  the  extension  of  pseudopodia  as  demonstrated  by  the 
plentiful  formation  of  pseudopodia  upon  fixation  with  osmic  acid; 
OH  ions  cause  an  increase  of  surface  tension  and  a  retraction  of 
pseudopodia.  If  we  imagine  a  bacterium  to  be  a  negatively  charged 
particle  which  gives  off  H  ions,  by  dissociation  it  will  lower  the  surface 
tension  at  the  presenting  point  of  the  ameba  and  occasion  the  appear- 
ance of  pseudopodia.  When  the  bacterium  is  surrounded,  there  is  an 
equalization  of  charge,  the  surface  tension  is  raised  and  the  pseudo- 
podium  is  retracted  with  the  bacterium.  L.  Hirschfeld  attributes 
the  positive  charge  of  amebse  to  the  excretion  of  CO2.  If  the 
metabolism  of  the  ameba  is  impaired,  the  formation  of  CO2,  and 
with  it  the  mobility  of  the  ameba,  are  diminished.  What  occurs  in 
the  case  of  amebse  may  be  applied  to  the  special  case  of  phagocytosis. 
It  was  the  phenomena  occurring  in  amebse  that  led  Elie  Metchni- 
KOFF  to  his  fundamental  studies  on  phagocytes,  scavenger  cells. 
Thus  he  names  such  white  blood  corpuscles  as  attack  by  taking  up 
and  digesting  microorganisms  entering  the  blood  stream.  They  are 
the  defending  army  of  the  organism,  and  according  to  E.  Metchni- 
KOFF,  the  most  important  weapon  in  the  fight  against  disease  germs. 


286 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


L.  Hirschfeld  is  supported  in  his  theory  by  the  statement  of  H. 
Bechhold*  that  lactic  acid  (H  ions)  increase  the  phagocytic  activ- 
ity of  leucocytes,  whereas  alkalis  (OH  ions)  are  without  such  effect. 

This  introduces  us  to  one  of  the  most  important  fields  as  yet  [al- 
most] untouched  by  colloid  investigation,  Chemotaxis,  the  experimental 
study  of  which  from  modern  viewpoints  ought  to  prove  most  promising. 

In  1884,  E.  Stahl  and  de  Bary  on  the  one  hand,  and  W.  Pfeffer 
on  the  other,  simultaneously  gave  their  attention  to  the  nature  of 
Chemotaxis.  They  studied  the  lower  monera,  plasmodia  of  mjrxo- 
mycetes  (slime  fungi)  bacteria,  flagellates,  wheel  animalcules,  the 
clustered  spores  of  algae,  the  spores  of  ferns,  mosses,  etc.  The  essence 
of  Chemotaxis  lies  in  the  attraction  of  these  unicellular  organisms 
by  certain  substances  (positive  Chemotaxis)  and  their  repulsion  by 
others  (negative  Chemotaxis),  while  other  substances  do  not  affect 
them  at  all.  If  for  instance  a  cane  sugar  solution  is  placed  in  a 
very  narrow  test  tube,  and  the  open  end  is  dipped  into  a  drop  of 
moss  spores,  the  latter  will  pass  into  the  tube,  attracted  by  the 
cane  sugar.  It  is  necessary  to  assume  some  such  chemotactic  re- 
lation between  eggs  and  spermatozoa,  especially  of  aquatic  animals, 
as  the  spermatazoa  discharged  into  the  water  are  attracted  by  the 
eggs.  We  owe  our  knowledge  of  the  chemotactic  action  of  leuco- 
cytes of  the  higher  animals  to  C.  A.  Pekelharing  and  especially  to 
Th.  Leber  who  gives  in  his  classical  work,  "Die  Entstehung  der 
Entzündung,"  a  wealth  of  experiments  in  which  the  most  varied 
substances  were  introduced  into  the  eyes  of  rabbits.  In  the  same 
field,  but  it  is  quite  obvious  independently,  he  was  followed  by  Mas- 
sart and  J.  Bürdet. 

We  reproduce  the  following  series  of  substances  with  a  chemotactic 
action  (after  A.  Gabritschewsky)  to  show  how  difficult  it  is  to  ex- 
plain the  existence  of  Chemotaxis  on  a  single  principle. 

Substances  Showing  Chemotaxis. 


Negative. 

Absent. 

Positive. 

10%  K  and  Na  salts 

Distilled  water 

1%  papayotin  (for  rabbits) 

l-io%  glycerin 

Carmin  powder 

Living  and  killed  cultures  of: 

Bile 

0.1  to  1%  K  and 

Bacillus  pyocyaneus 

10%  alcohol  _ 

Na  salts 

Bacillus  prodigiosus 

Chloroform  in  aqueous 

Phenol 

Bacillus  of  anthrax 

solution 

1%  antipyrin 

Bacillus  of  typhoid 

0.5%  quinine  solution 

1%  phloridzin 

Bacillus  of  hog  erysipelas 

0.1  to  10%  lactic  acid 

1%  papayotin  (for 

All    the    bacteria    that    have 

Jequirity 

frogs) 

been  studied  excepting  the 

Sterile  culture  of  chicken 

1%  glycogen 

bacilli  of  chicken  cholera 

cholera  bacilli 

1%  peptone 
Bouillon 
Aqueous  humor 

Blood 

THE  MOVEMENTS  OF  ORGANISMS  287 

It  may  be  concluded  ffbm  this  that  a  substance  may  be  neutral  for 
certain  leucocytes  and  positively  chemotactic  for  others  (papayotin), 
and  that  the  chemotactic  relation  may  vary  with  concentration 
(K  and  Na  salts;  with  reference  to  lactic  acid  and  quinine  solutions 
see  pp.  286-288). 

What  relation  does  all  this  bear  to  the  theory  of  changing  surface 
tension?  Some  data  are  in  its  favor.  The  very  first  observation  of 
E.  Stahl  on  the  plasmodia  of  sethalium  septicum  of  tanner's  bark  gives 
a  decided  impression  that  a  surface  phenomenon  is  involved.  When 
he  brought  such  a  plasmocUum  clinging  to  the  internal  surface  of  a  glass 
in  contact  with  pure  water  by  introducmg  the  water  from  below,  the 
Plasmodium  spread  out  uniformly;  if  he  introduced  tannic  acid,  it  trav- 
eled downwards;  and  on  the  addition  of  from  1/4  to  1/2  per  cent  sugar 
solution,  it  traveled  upwards.  It  is  just  this  action  of  tannic  acid 
which  tans  the  surface  of  protoplasmic  mucus  and  the  phenomenon 
of  spreading  out  in  pure  water  that  point  to  surface  forces.  They  are 
also  suggested  by  the  observation  of  Ranvier,  according  to  whom 
leucocytes  spread  out  more,  the  larger  the  surface  development  of  the 
given  body  (better  on  rough  than  on  smooth  surfaces  and  especially 
well  upon  elder  pith).  On  the  other  hand,  we  recognize  from  what 
has  been  said  that  the  theory  which  attributes  the  decrease  in 
surface  tension  to  an  electrical  charge  does  not  suffice  for  the  ex- 
planation of  all  phenomena.  An  intensely  positive  chemotactic 
action  is  possessed  not  only  by  bacteria,  but  also  by  extracts  and 
proteins  obtained  from  them.  The  chemotactic  experiments  under- 
taken on  the  bodies  of  higher  animals  (eye,  pleura,  etc.)  do  not 
justify  a  physico-chemical  explanation,  because  in  this  instance  two 
factors  coexist.  The  substance  itself  may  act  chemotactically; 
on  the  other  hand,  it  may  be  inactive  yet  cause  a  necrosis  of  the 
adjoining  tissue,  which  then  becomes  chemotactic  and  simulates 
activity  on  the  part  of  the  substances  under  investigation.  [Else- 
where (p.  234)  reference  has  been  made  to  the  observations  of 
A.  B.  Macallum.  His  monograph  "  Surface  Tension  and  Vital 
Phenomena,"  No.  8  Physiological  Series,  University  of  Toronto 
Studies,  1912,  includes  a  bibliography.     Tr.] 

Possibly  the  very  original  "Quantitative  Studies  on  Phagocytosis" 
of  H.  J.  Hamburger  and  Hekma*  will  permit  conclusions  concern- 
ing the  causes  of  the  protoplasmic  movements  of  leucocytes,  when  a 
method  shall  have  been  discovered  for  measuring  the  surface  tension 
of  protoplasm  against  water  and  salt  solution.  Even  now  it  may  be 
recognized  from  these  studies  that  the  causes  of  movement  are  quite 
complicated  since  it  has  been  shown  that  the  calcium  ion  has  an 
entirely  specific  action  in  stimulating  phagocytosis.     If  such  action 


288  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

were  due  merely  to  the  electric  charge  possessed  by  Ca  as  a  divalent 
ion,  we  would  expect  the  same  effect  from  barium,  strontium  and 
magnesium;  this  however  is  not  the  case. 

Especially  noteworthy  is  the  fact,  only  recently  studied  by  G. 
Denys  and  Leclef,  Weight  and  his  pupils,  and  Neufeld  among 
others,  that  leucocytes  are  stimulated  to  the  phagocytosis  of  certain 
bacteria  only  by  the  presence  of  serum,  and  that,  on  the  one  hand,  the 
intensity  of  the  phagocytosis  is  dependent  upon  the  virulence  of  the 
bacteria,  and,  on  the  other,  upon  certain  properties  of  the  serum, 
closely  related  to  those  which  determine  immunity. 

To  the  colloid  chemist,  it  is  of  importance  to  determine  whether 
the  general  colloid  properties  of  serum  play  a  role  in  phagocytosis, 
and  whether  the  serum  may  be  replaced  by  other  colloids.  H. 
Bechhold*  showed  that  egg  albumen,  which  stands  nearest  to  serum 
in  respect  to  its  colloid  properties,  caused  no  phagocytosis,  whereas 
Witte's  peptone,  a  markedly  broken  down  protein,  has  such  an 
action. 

In  the  case  of  Chemotaxis,  as  in  the  case  of  phagocytosis  under 
the  influence  of  opsonins  (or  certain  hypothetical  irritants  which 
increase  the  appetite  of  leucocytes)  only  comprehensive  quantitative 
experiments  will  yield  material  utilizable  for  the  development  of 
a  physico-chemical  theory  by  the  colloid  chemist.  Although,  for 
instance,  quinine  is  regarded  as  a  substance  which  inhibits  phago- 
cytosis, M.  Neisser  and  Guerrini  *  have  shown  that  in  minimal 
doses  it  increased  the  appetite  of  leucocytes. 

It  may  be  said  in  conclusion  that  the  surface  tension  of  leucocytes 
in  relation  to  the  surrounding  medium  (serum)  must  be  very  low. 
On  page  16,  we  saw  what  force  is  necessary  to  change  the  form  of  such 
small  bodies  (leucocytes  have  an  average  diameter  of  from  6  to  8  /x). 
If  we  recall  what  changes  in  surface  tension  a  leucocyte  may  undergo 
in  phagocytosis,  and  the  very  great  changes  in  shape  suffered  in 
traversing  the  tissues,  we  are  forced  to  ascribe  to  them  a  very  low 
surface  tension,  much  lower  than  that  possessed,  e.g.,  by  red  blood 
corpuscles. 

The  Movements  of  Higher  Organisms. 

The  movements  of  higher  organisms  are  controlled  by  the  nerves 
and  accomplished  by  the  muscles.  In  the  present  state  of  our 
knowledge  and  in  the  limits  of  this  book  we  can  only  consider  this 
question:  From  what  physical  and  chemical  processes  does  muscle 
contraction  result?  For  this  purpose  we  shall  first  consider  the  muscle 
as  a  colloid  system  and  endeavor  to  gain  an  idea  how  a  contraction 
occurs. 


THE,  MOVEMENTS  OF  ORGANISMS  289 

Muscle  as  a  Colloid  System. 

In  the  case  of  higher  mammals,  muscles  constitute  approximately 
43  per  cent  of  the  entire  body.  Since  they  have  a  greater  range  of 
swelling  than  all  the  other  organs  (see  p.  219),  besides  their  usual 
function  as  a  water  reservoir,  they  are  of  great  importance. 

As  regards  swelling,  they  behave  very  much  like  fibrin  or  gelatin. 
It  was  formerly  believed  that  the  circumstances  of  sweUing  in  muscle, 
which  were  at  first  chiefly  studied  in  the  case  of  frog  muscle,  could 
be  explained  by  osmosis,  but  the  quantitative  studies  of  J.  Loeb,* 
followed  later  by  A.  Durig,  C.  E.  Overton  *  and  R.  W.  Webster, 
showed  that  no  satisfactory  solution  could  be  thus  obtained.  If 
the  osmotic  conditions  alone  were  determinative,  the  muscle  should 
retain  its  water  in  isotonic  solutions,  shrinking  in  hypertonic  and 
swelling  in  hypotonic  solutions.  But  this  is  not  by  any  means  the 
case,  since  there  is  a  material  difference  between  solutions  of  electro- 
lytes and  of  nonelectrolytes.  Whereas  neutral  salts  greatty  diminish 
the  swelling  produced  by  acids  and  alkalis,  this  property  is  not  pos- 
sessed by  nonelectrolytes  (cane  sugar,  ethyl  alcohol,  methyl  alcohol, 
urea  and  glycerin).  Even  the  supposition  of  a  lipoid  membrane 
does  not  explain  the  phenomena,  since  cane  sugar  is  as  insoluble 
in  lipoids  as  are  most  of  the  neutral  salts. 

As  early  as  1901,  A.  Durig  concluded  from  his  investigations  with 
whole  frogs  that  the  laws  which  are  invoked  in  osmotic  processes 
alone  are  inadequate;  in  this  case  muscles  are  chiefly  concerned  in 
the  absorption  and  relinquishment  of  water.  Martin  H.  Fischer* 
was  the  first  to  direct  attention  to  the  fact  that  for  dead  muscle, 
qualitatively  and  to  some  extent  quantitatively,  similar  laws  gov- 
erned the  taking  up  and  the  relinquishment  of  water  as  governed 
unorganized  colloids  capable  of  swelling. 

To  summarize  his  results  briefly:  muscles  swell  more  in  acids  and 
in  alkalis  than  in  water,  and  indeed,  in  hydrochloric  acid,  nitric  acid  > 
acetic  acid  >  sulphuric  acid.  The  maximum  amount  of  water  that  a 
muscle  can  absorb  under  the  circumstances  is  about  246  per  cent  of 
the  original  muscle  weight,  or  13  times  the  dried  muscle  substance. 
It  therefore  possesses,  it  is  true,  a  smaller  swelling  capacity  than 
gelatin  which  can  take  up  from  15  to  25  times,  or  fibrin  which  takes 
up  upon  solution  30  to  40  times,  its  dried  weight. 

The  absorption  and  relinquishment  of  water  by  muscle  is  a  re- 
versible process,  yet  M.  H.  Fischer  emphasizes  the  fact  that 
during  the  time  of  his  experiments  no  complete  reversibility  was 
observed,  that  "every  change  of  condition  left  its  permanent 
results." 


290  COLLOIDS  IN  BIOLOGY  AND   MEDICINE 

Salts  diminish  the  swelhng  of  muscle  in  acids  and  alkalis  in  a  way 
similar  to  the  case  of  fibrin  and  gelatin,  though  not  so  obviously. 

There  is,  indeed,  a  very  important  difference  between  dead  and 
living  muscle:  the  swelling  of  dead  muscle  in  distilled  water,  for 
instance,  is  brought  about  by  the  formation  of  lactic  acid,  which  sets 
in  within  a  few  minutes.  If  this  were  not  the  case,  a  living  frog  would 
swell  up  as  much  in  fresh  water  as  a  dead  one.'^  According  to  M.  H. 
Fischer,  a  dead  muscle  retains  its  form  in  a  0.7  per  cent  NaCl  solu- 
tion, not  because  the  same  osmotic  pressure  exists  inside  and  outside 
the  cell,  but  because  the  concentration  of  the  NaCl  solution  is  just 
sufficient  to  overcome  the  action  of  the  acids  formed  in  the  excised 
muscle.  We  must  again  point  out  here  that  the  experiments  of  W. 
BiLTZ  and  A.  von  Vegesack  *  show  that  if  colloids  are  present  in  a 
medium,  the  presence  of  isotonicity  does  not  by  any  means  permit  us 
to  infer  that  equal  osmotic  pressures  exist. 

Against  M.  H.  Fischer's  experiments,  the  objection  has  been 
raised  that  dead  muscle  possesses  no  semipermeable  membrane,  so 
that  its  swelling  follows  laws  similar  to  those  of  fibrin,  etc.  In  living 
muscle,  however,  semipermeability  exists;  on  this  account  the  re- 
sults of  M.  H.  Fischer  cannot  be  transferred  to  living  muscle. 
There  are  also  certain  discrepancies  in  respect  to  some  nonelectro- 
lytes;  thus,  for  instance,  dead  muscle  does  not  swell  up  in  isotonic 
sugar  solution;  this  does  not  accord  with  Fischer's  theory.  [Sugar 
has  a  specific  dehydrating  action.     Tr.] 

The  studies  of  E.  B.  Meigs  *  have  illuminated  these  discrepancies; 
they  showed  a  definite  difference  between  smooth  and  striated  muscles. 
Smooth  muscles  are  involuntary  and  occur  in  automatically  acting 
organs  (intestines,  urinary  bladder,  iris,  etc.),  and  especially  widely 
distributed  among  the  lower  animals.  They  contract  much  more 
slowly  than  striated  muscles.  E.  B.  Meigs  concludes  that  smooth 
muscle  is  not  surrounded  by  a  semipermeable  membrane,  in  other 
words,  osmosis  is  not  a  factor,  but  that  they  behave  toward  electro- 
lytes like  any  hydrophile  colloid,  fibrin  or  gelatin,  with  reference  to 
change  in  volume. 

The  behavior  of  striated  muscles  is  quite  different.  To  understand 
it  we  must  briefly  recall  their  histology.  Muscles  consist  of  bundles 
of  fibrils,  longitudinal  fibers  which  are  surrounded  by  a  connective 
tissue  sheath.  Each  fibril,  that  is,  every  minute  fiber,  is  surrounded 
by  a  membrane,  the  sarcolemma,  and  is  bathed  in  a  fluid  substance,  the 
sarcoplasm.  The  individual  fibrils  are  striated  at  right  angles  to  their 
axes.  The  striations  appear  microscropically  as  alternating  dark  and 
bright  zones;  while  the  latter  are  isotropic,  the  dark  striations  are 
doubly  diffractive,  anisotropic  (see  Fig.  49). 

1  [If  the  circulation  of  a  living  frog  is  impeded  so  that  local  acidosis  develops, 
local  swelling  also  develops.     Tr.] 


THE  MOVEMENTS  OF  ORGANISMS 


291 


Fig.  49.  Striated 
muscle  fiber. 
(Stöhr.) 


E,  B.  Meigs  *  studied  the  rate  at  which  fresh  muscles  in- 
creased their  weight  in  water  and  in  salt  solutions.  He  concluded 
from  his  study  that  the  weight  increase  is  the  result  of  two  processes : 
At  first,  water  is  osmotically  taken  up  by  the  sarcoplasm  of  the  fresh 
(still  irritable)  muscle;  after  the  muscle  is  dead,  lactic  acid  forms, 
the  semipermeable  membrane  of  the  fibrils  (the  sarcolemma)  becomes 
permeable  and  now  the  fibrils  swell  up  at  the  ex- 
pense of  the  sarcoplasm  fluid  and  are  thus  'short- 
ened; this  is  evidenced  by  rigor  mortis  (0.  von 
FÜRTH  and  Lenk).  The  proteins  become  co- 
agulated through  the  accumulation  of  acid;  this 
especially  induces  a  shrinking  and  thus  a  relaxa- 
tion of  rigor  mortis.  By  this  experimentally  es- 
tabHshed  explanation  0.  von  Fürth  and  Lenk 
have  cleared  away  an  old  fallacy  that  the  onset 
of  coagulation  induced  rigor  mortis.  By  artifi- 
cial fatigue  {e.g.,  electrical  stimulation  of  an 
excised  frog's  muscle)  the  accumulation  of  acid 
and  the  consequent  swelling  of  muscle  in  dilute 
salt  solution  is  much  hastened  (C.  Schwarz  *).  It  is  a  well-kno^vn 
fact,  moreover,  that  after  great  muscular  exertion  (forced  marches, 
convulsions,  hunted  prey),  rigor  mortis  sets  in  sooner  than  when 
death  overtakes  a  rested  organism. 

When  rigor  mortis  disappears  striated  muscle  behaves  like  an 
hydrophile  colloid,  whose  swelling  and  shrinking  are  unhindered  by 
semipermeable  membranes. 

A  further  study  of  E.  B.  Meigs*  is  concerned  with  the  nature  of 
the  semipermeable  membrane  of  a  fibril.  It  tends  to  show  that  the 
latter  consists  of  calcium  phosphate.  Collodion  membranes  impreg- 
nated with  calcium  phosphate  proved  impermeable  for  salts,  sugar 
and  amino  acids,  but  were  somewhat  permeable  for  glycerin  and  urea 
and  easily  permeable  for  ethyl  alcohol.  They  were  moderately  perme- 
able for  potassium  chlorid  as  was  to  be  expected.  The  predication  of 
a  semipermeable  layer  of  calcium  phosphate  explains  two  facts  very 
well:  1.  The  suspension  of  the  semipermeabiHty  of  muscle  after 
death  (the  accumulation  of  lactic  acid  destroys  the  membranes)  and 
2.  the  importance  of  calcium  for  the  maintenance  of  semipermea- 
biHty in  living  muscle;  since  the  layer  of  calcium  phosphate  is  de- 
stroyed in  a  neutral  lime-free  solution. 

A  unique  observation  was  made  by  M.  H.  Fischer  and  P.  Jensen  * 
upon  the  water  in  muscle.  They  put  the  gastrocnemius  of  frogs  into 
narrow  glass  tubes,  cooled  them  do\^^l  to  —76°  in  a  mixture  of 
ether  and  solid  CO2,  and  followed  the  curve  of  cooling  wdth  a  needle- 


292  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

shaped  thermocouple.  With  a  muscle  of  average  size,  the  phenomena 
are  about  as  follows:  Within  3  or  4  minutes  there  is  a  very  slight 
cooling;  within  another  8  or  10  minutes  the  water  freezes  in  the 
muscle  and  further  cooling  occurs.  This  part  of  the  curve  is  quite 
characteristic  for  the  fixation  of  water.  It  should  fall  more  steeply 
than  the  curve  of  control  water  or  of  physiological  salt  solution.  If 
it  is  less  steep,  it  is  "a  sign  that  there  is  some  process  in  the  colloid 
structures  which  hberates  heat."  ^  In  H.  W.  Fischer's  and  P.  Jen- 
sen's investigations,  it  was  shown  that  it  is  necessary  to  distinguish 
two  kinds  of  water  fixation  in  freezing  muscle.  ''After  or  during 
death  by  freezing,  there  occurs  a  phenomenon  by  which  water  is  fixed 
in  some  unknown  way  and  by  which  it  is  again  liberated  at  lower  tem- 
peratures," and  indeed  the  amount  of  water  fixed  in  a  muscle  in- 
creases with  the  amount  of  disturbance  (whether  frozen  once,  twice, 
heated  to  100°  or  boiled).  In  this  case,  also,  it  is  seen  that  two  kinds 
of  water  fixation  exist. 

The  relation  between  cooling  and  the  death  of  muscle  by  freezing 
is  very  interesting.  The  degree  of  "vitality^'  was  measured  by  the 
lifting  capacity  of  a  muscle  in  response  to  stimulation.  It  was 
shown  that  cooling  the  muscle  to  the  point  of  freezing,  and  even 
freezing  out  the  water  to  a  certain  extent,  did  no  harm,  but  if  the 
muscle  was  cooled  1.5°  C.  more,  it  died.  The  inferior  thermal  margin 
between  life  and  death  of  muscle  is,  therefore,  only  1.5°  C.  wide. 

The  normal  state  of  swelling  in  muscle  is  conditioned  by  a  normal 
content  of  electrolyies.  This  may  vary  greatly  for  different  classes 
of  animals;  for  instance,  according  to  J.  Katz,  the  striated  muscle 
fibers  of  the  dog  contain  3.5  times  as  much  K,  and  those  of  the  pike 
14  times  as  much  K,  as  Na.  For  the  same  species  it  appears  to  be 
uniform  at  the  same  age. 

A  remarkable  fact  regarding  muscles  is  their  high  potassium  content, 
which  is  closely  associated  with  their  capacity  to  functionate.  [See 
Macallum,  also  Burridge.  Tr.]  For  a  normal  swelUng,  the  iso- 
tonicity  of  the  surrounding  solution  appears  to  be  of  much  less 
importance  than  a  definite  electrolyte  content.  This  follows  from 
experiments  of  E.  M.  Widmark,*  according  to  which  even  10  milli- 
mols  CaCl2  in  the  surrounding  solution  produced  a  loss  of  weight 
amounting  to  36  per  cent  (in  the  spUt  muscle  fibers). 

Muscle  Function. 

Every  stimulus,   whether  of  thermal,   mechanical,   electrical  or 
chemical  nature  causes  an  irritation  in  the  living  muscle  which  is 
manifested  by  a  contraction.     R.  Höber,*  whose  investigations  we 
^  The  mathematical  basia  for  this  is  given  in  the  original  work. 


THE  MOVEMENTS  OF  ORGANISMS  293 

shall  now  consider,  is  primarily  responsible  for  the  electrochemical 
theory  of  this  irritability. 

For  our  consideration,  two  electrical  phenomena  of  muscle  are 
important:  In  activity,  that  is  during  the  contraction  of  muscle, 
electric  currents  (action  currents)  develop;  the  stimulated  point  in 
the  muscle  becomes  negative  in  relation  to  the  remaining  fibers  which 
are  at  rest.  The  same  thing  holds  true  for  nerves  in  which  no  external 
sign  of  activity  is  discoverable. 

If,  in  an  excised  mollusc  muscle,  an  injured  point  is  united  to  an 
uninjured  point  of  the  mantle  by  a  wire  having  a  galvanometer  in 
its  circuit,  the  cut  surface  is  negative  and  the  mantle  surface  is  posi- 
tive. The  same  electrical  phenomena  are  observed  in  a  resting  nerve. 
This  is  called  the  current  of  rest,  or,  according  to  H.  Herman,  the  de- 
marcation current  (Herman  calls  the  demarcation  surfaces  the  interface 
between  the  injured,  dead,  and  the  uninjured,  Hving,  substance). 

Evidently  action  current  and  current  of  rest  are  due  to  the  same 
cause.  In  his  textbook,  R.  A.  A.  Tigerstedt  states  the  phenomenon 
as  follows :  In  muscle  as  in  nerve,  a  stimulated  point,  or  one  which  is 
injured  in  any  way,  is  negative  electrically  to  every  other  point  which 
at  that  time  happens  to  be  at  rest  or  uninjured. 

Let  us  consider  how  we  may  explain  the  direction  and  magnitude 
of  different  potentials  which  occur  when  muscle  contracts.  Elec- 
trical differences  in  potential  arise  on  every  interface  between  an 
electrolyte  and  a  pure  solvent  or  one  containing  less  electrolytes. 
The  simplest  case  is  when  an  acid,  e.g.,  HCl,  is  Hmited  by  pure  water  — 
then  the  more  mobile  positive  H  ion  will  rapidly  advance  and  give  a 
positive  charge  to  the  water  while  the  acid  is  negatively  charged  by 
the  more  slowly  moving  negative  CI  ion.  This  applies  to  muscle, 
for  lactic  acid  arises  at  the  point  stimulated  or  injured. 

The  electromotive  forces  which  are  derived  from  a  circuit  of  acids 
and  water  or  crystalloid  electrolytes  are  much  smaller  than  we 
observe  in  muscle. 

Wo.  Pauli  invokes  the  colloidal  properties  of  the  protein  ions  in 
explaining  the  high  electric  tension  which  we  obtain  in  muscle  or  even 
in  the  electric  organ  of  the  torpedo. 

Protein  in  general  contains  an  amino  acid  with  many  NH2  and 
COOH  groups.  Let  us  illustrate  the  development  of  electromotive 
forces  by  the  following  diagram  in  which  R  represents  the  protein 
radicle  and  L  the  lactic  acid  radicle: 

OHCO  .    .  NH2  +  LH   OHCO  •    •  NH3   L 

OHCO.  |<  .NH2  +  LH  =  0HC0.  P-NHg-fL 

OHCO-    .NH2  +  LH   OHCO.    .NH3   L 


294  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  difficultly  mobile  colloidal  acid-protein  ion  immediately  becomes 
positively  charged  at  the  surface  of  a  neutral  medium,  and  should  it 
touch  an  acid  medium  its  positive  charge  is  raised  and  at  the  same 
time  the  acid  field  becomes  more  negative  as  the  following  diagram 
indicates : 


OH .  CO  ■ 

.NH2 

OH . CO . 

Tl'T% 

OH .  CO . 

R 

.  NH2  +  LH  = 

+ 

=  OH . CO ■ 

|<  •  NH2  H  +  L 

OH .  CO . 

-ML.mJ 

.NH2 

OH . CO ■ 

.          .  NH2 

for  the  H  ion  moves  faster  than  the  L  ion.  Measurements  of  series 
consisting  of  acids  and  acid  albumin  couples  yielded  potentials  quite 
large  enough  to  account  for  action  currents. 

The  development  of  such  diffusion  potentials  in  muscle  would  not 
be  possible  if  the  fibrils  were  not  quite  poor  in  salt  and  the  sarcoplasm 
quite  rich  in  salt.  Since  both  the  fluid  and  the  fibrillar  portions 
contain  protein  (see  Bottozzi  and  his  school)  a  couple  consisting  of 
acid  albumin/acid/acid  albumin  yields  no  current.  The  current  is 
reestablished  through  electrolytic  dissociation  of  the  acid  albumin 
due  to  the  salt  in  the  sarcoplasm  (see  p.  292).  If  such  couples  are 
placed  in  series  considerable  electric  tension  (voltage)  may  be  ob- 
tained. 

These  results  are  in  agreement  with  the  fact  that  the  normal 
properties  of  muscle  are  conditioned  by  definite  states  of  swelling  and 
electrolyte  content. 

If  frogs'  muscles  are  placed  in  an  isotonic  solution  of  cane  sugar 
or  other  nonelectrolyte  (mannit,  asparagin,  etc.),  they  lose  their 
irritabihty  (e.g.,  for  the  induced  current)  but  retain  their  volume; 
they  do  not  swell  as  in  distilled  water  in  which  the  irritability  is 
likewise  suspended.  The  ability  to  contract  is  restored  by  Na  ions 
(about  0.07  per  cent  NaCl)  (C.  E.  Overton)  as  well  as  by  Li  ions; 
but  it  is  not  restored  at  all  by  K  ions.  The  irritability  is  also  sus- 
pended by  isotonic  potassium  and  rubidium  salts.  If  the  anions 
and  cations  are  arranged  in  accordance  to  the  extent  with  which 
they  interfere  with  irritability,  we  obtain  lyotropic  series  similar  to 
those  which  we  discovered  for  the  salting  out  of  colloids  (see  pp.  80 
to  83);  according  to  R.  Höber,*  C.  E.  Overton*  and  Schwarz,* 
they  are  as  follows: 

inhibitory:  K  >  Rb  >  Cs  >  Na,  Li 

inhibitory:   tartrate,  SO4  >  acetate  >  CI  >  Br,N03  >  I  >  SCN. 

If  an  uninjured  frog's  muscle  is  dipped  into  an  isotonic  solution 
of  a  neutral  salt  and  the  part  so  treated  is  united  with  another  part  of 


THE  MOVEMENTS  OF  ORGANISMS  295 

muscle  l?y  a  wire,  we  obtain  a  current  of  rest  whose  strength  and  direc- 
tion depends  on  the  nature  of  the  neutral  salt.  [The  study  of  these  cur- 
rents of  action  in  the  heart  muscle  has  been  elaborated  into  the  science 
of  electrocardiography,  I  know  of  no  attempt  to  associate  electro- 
cardiographic curves  with  changes  in  the  colloids  of  the  heart  muscle 
in  response  to  salts.  Tr.]  If  the  anions  and  cations  are  arranged 
according  to  their  action  on  this  current  of  rest  (see  R.  Höber  and 
Waldenberg  *),  we  obtain  series  similar  to  the  above.  Since  we 
have  previously  seen  that  the  salting  out  of  protein,  the  swelling  and 
shrinking  of  gelatin  and  fibrin  (which  means  the  ionization)  occur  in 
similar  lyotropic  series,  R.  Höber  concludes  that  the  normal  irrita- 
bility of  muscle  is  dependent  upon  a  definite  condition  of  solution  or 
swelling  of  its  protoplasmic  colloids;  increased  solution  or  precipitation 
of  the  colloids  leads  to  loss  of  irritability.  J.  Loeb  and  R.  Beutner 
are  of  the  opinion  that  the  current  of  inactive  muscle  due  to  salt 
(as  well  as  the  currents  rising  in  plants  because  of  an  injury  to  some 
part)  bears  no  direct  relation  to  the  condition  of  swelling  of  the 
plasma  colloids,^  but  is  due  to  a  hpoid  membrane  on  the  surface  of 
the  muscle  or  its  constituent  elements.  The  variation  in  activity  of 
the  salts  chosen  (NaCl,  KCl,  etc.)  is  due  to  their  different  threshold 
of  solubility  in  the  hpoid  membrane. 

R.  HÖBER  correctly  emphasized  that  for  such  questions  of  physio- 
logical function  we  need  consider  only  those  influences  which  are 
reversible.  Substances  causing  a  more  or  less  irreversible  change  by 
means  of  aromatic  anions  require  no  further  consideration  here. 

The  dependence  of  the  irritab  hty  of  muscle  upon,  and  its  relation 
to,  the  condition  of  the  organ  colloids  are  not  unique.  Examples  of 
other  organ  functions  were  studied  by  R,  S.  Lillie  *^  (movement  of 
the  cilia  of  the  larvae  of  marine  annehds)  and  by  R.  Höber  *^  (the 
movement  of  the  cihated  epithelium  of  the  frog) . 

The  movements  of  cilia  above  mentioned  cease  upon  the  addition 
of  various  salts:  in  fact,  of  the  alkali  salts,  Li  salts  are  the  most 
harmful.  In  hemolysis  and  in  the  diminution  of  the  movement  of 
ciha,  the  anion  series  shows  an  order  the  reverse  of  that  for  the 
diminution  of  muscle  irritability,  which  means  that  the  swelling  of 
blood  corpuscles  and  muscle  are  affected  in  an  opposite  way.  Such 
well-known  hemolytic  agents  as  saponin,  solanin,  taurocholic  acid, 
glycochohc  acid  and  sodium  oleate  diminish  the  irritability  of  muscle 
in  an  irreversible  manner;  they  evidently  damage  the  lipoid  plasma 
pelhcle  (R.  Höber  *ii). 

1  We  must  forego  further  discussion  of  the  extremely  interesting  results  of 
J.  Loeb  and  R.  Beutner  since  they  have  no  direct  bearing  on  colloids. 


296  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  accompanying  table  (in  part  after  R.  Höber)  gives  at  a  glance 
the  action  of  the  various  alkaline  salts,  and  parallel  with  it  the  extent 
to  which  such  salts  salt  out  hydrophile  colloids. 

The  question  now  arises,  What  are  the  colloid-chemical  changes 
which  occur  as  the  result  of  stimulation  and  bring  about  the  change  in 
the  shape  of  the  muscle  f 

We  know  from  the  investigations  of  G.  Jappelli  and  D'Errico  as 
well  as  of  G.  Buglia,  that  muscle  absorbs  water  when  it  contracts 
(fatigue).  This  is  not  surprising,  since  acids  are  formed  which 
favor  swelhng  (see  p.  267).  According  to  the  conception  of  E. 
Pribram,  the  formation  of  acids  and  the  contraction  of  muscle  are 
closely  associated.  Even  Th.  W.  Engelmann  had  already  drawn 
the  conclusion  that  during  contraction,  water  passes  from  the  iso- 
tropic water-rich  layer  of  the  striated  muscle  into  the  anisotropic 
water-poor  layer,  which  swells.  This  is  due  to  the  transfer  of  acids 
from  the  sarcoplasm  where  lactic  acid  is  created  by  stimulation. 
Water  flows  from  the  blood  and  the  lymph  into  the  isotropic  layer,  so 
that  as  a  result  of  the  contraction,  the  entire  muscle  is  richer  in 
water.  We  must  picture  of  the  shifting  of  fluid  within  the  fibrils  as 
occurring  in  such  a  way  that  the  anisotropic  layer,  which,  according  to 
MUNCH,  is  spirally  arranged,  can  expand  only  from  side  to  side  when  it 
swells  (at  the  expense  of  the  isotropic  layer).  This  causes  a  trans- 
verse thickening  of  the  muscle  fibers  and  a  shortening  in  length,  a 
contraction.  If  the  lactic  acid  in  the  living  muscle  is  consumed  or 
otherwise  neutralized  the  process  is  reversed  and  the  muscle  regains 
repose. 

Streitmann  and  M.  H.  Fischer  constructed  from  catgut  a  working 
model  of  muscular  contraction.  The  catgut  strands  represented  the 
anisotropic  substance  and  the  sarcoplasm  was  replaced  by  water, 
acids,  and  salt  solutions. 

For  the  sake  of  completeness,  we  shall  refer  to  one  other  theory 
which  is  by  no  means  as  well  estabhshed  experimentally  as  the  one 
described.  Bernstein  first  suggested  the  idea  that  muscular  con- 
traction was  associated  with  changes  in  surface  tension.  As  has  been 
mentioned  previously,  muscle  is  characterized  by  an  especially  high 
content  of  potassium.  From  the  researches  of  A.  B.  Macallum  we 
are  compelled  to  assume  that  it  has  a  most  important  function  dur- 
ing contraction. 

In  contractile  tissues  (muscles  of  frogs,  lobsters,  beetles,  etc.), 
according  to  A.  B.  Macallum  *  and  his  pupils,  the  potassium  seems 
to  be  localized  in  the  dark  zones  of  the  resting  muscle  fibrils,  especially 
at  their  surfaces.  From  this,  A.  B.  Macallum  concludes  that  the 
surface  tension  must  be  lowered  in  these  zones.     With  the  contrac- 


THE  MOVEMENTS  OF  ORGANISMS 


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298  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

tion  there  occurs  a  change  in  the  distribution  of  potassium.  There  is 
thus  associated  with  each  contraction  and  return  to  rest  a  shifting 
of  potassium,  a  shifting  of  the  swelling  and  a  change  in  the  surface 
tension.  We  may  leave  undecided  which  phenomena  are  primary  and 
which  are  secondary.  A.  B.  Macallum  calls  attention  to  the  fact 
that  next  to  hydrogen,  the  potassium  ion  has  a  greater  mobility  than 
any  other  cation,  and  attributes  to  this  the  rapidity  of  the  change  in 
surface  tension,  and  the  rapid  contractility  of  muscle. 

The  efficiency  of  muscle  depends  to  a  marked  extent  upon  its 
content  of  water  (see  J.  Demoor  and  Philippson  *)  and  may  be 
influenced  by  extreme  dehydration  (shrinking).  Such  extreme 
shrinking  may  be  brought  about  by  the  introduction  of  concentrated 
salt  solutions  or  glycerin  as  well  as  of  numerous  poisons  (especially 
veratrin)  (see  Santessow*  and  Gregor*).  Under  these  circum- 
stances the  muscle  is  shortened,  as  when  it  is  very  much  fatigued,  by 
tetanic  contractions. 

Likewise,  by  unsuitable  nutrition,  which  results  in  a  greatly 
swollen  state,  the  efficiency  of  the  muscle  may  be  depressed. 
TsuBOi  brought  about  such  a  swelling  in  rabbits  which  were  fed 
entirely  on  potatoes.  The  water  content  of  their  muscles  was  from  2 
to  7  per  cent  higher  than  normal.  Potatoes  are  especially  rich  in 
potassium,  and  in  this  connection  it  is  natural  to  think  of  the  swelling 
which  potassium  salts  also  cause  in  gelatin  and  fibrin,  and  of  the 
influence  of  the  K  ion  on  the  depression  of  muscle  irritability,  dis- 
covered by  R.  Höber. 

[W.  BuRRiDGE  associates  the  occurrence  of  fatigue  phenomena  with 
the  accumulation  of  K  ions  in  muscle  and  in  blood.  Adrenin  an- 
tagonizes K  salts. 

In  their  Croonian  Lecture  on  "  The  Respiratory  Process  in  Muscle 
and  the  Nature  of  Muscular  Motion,"  W.  M.  Fletcher  and  F.  G. 
Hopkins  come  to  the  conclusion  that  lactic  acid  is  not  a  toxic  product 
but  an  essential  agent  in  the  muscular  contractions.  Its  free  H  ions 
in  the  presence  of  colloidal  fibrils  cause  an  increase  in  tension  in  the 
fiber,  either  by  increasing  the  muscular  tension  along  the  longitudinal 
surfaces  or  by  the  process  of  imbibition.  They  studied  the  effect  of 
oxygen  on  muscle  and  found  that  it  not  only  delays  the  stiffening  of 
muscle  but  may  altogether  inhibit  its  onset.  A  muscle  forced  by 
stimulation  to  stiffening  may  be  recalled  again  by  oxygen  to  its  pre- 
vious flaccidity.  It  was  shown  that  immersion  of  a  fatigued  muscle 
in  oxygen  restored  the  osmotic  properties  to  those  of  resting  muscle. 
Fatigued  muscle  contains  more  lactic  acid  than  resting  muscle,  and 
a  fatigued  muscle,  after  resting  in  an  oxygen  atmosphere,  subsequently 
contained  less  lactic  acid.  A.  V.  Hill  and  Parnas  from  studies  of 
heat  production  in  contracting  muscle  conclude  that  the  combustion 


THE  MOVEMENTS  OF  ORGANISMS  298a 

of  the  lactic  acid  to  C 02' furnishes  the  heat  which  restores  some  of 
the  lactic  acid  to  its  metabolic  som'ce,  thus  removing  the  acid  ions 
from  the  colloidal  fibrils,  so  that  they  may  return  to  their  former 
tension  —  the  tension  of  rest  or  relaxation  —  and  possessed  of  their 
inherent  potential.  Proceedings  of  the  Royal  Society  of  London, 
Series  B  69,  Vol.  LXXXIX,  p.  444  et  seq. 

Contraction  of  the  heart  is  a  special  instance  of  muscular  function 
upon  which  considerable  light  has  been  thrown  by  the  studies  of 

W.  BURRIDGE. 

He  views  excitation  as  a  coagulative  change  induced  through  the 
formation  of  a  calcium  compound.  According  to  MacDonald  this 
coagulative  change  is  accompanied  by  a  release  into  aqueous  solution 
of  previously  adsorbed  K  salts  which  now  confer  a  positive  charge  on 
the  colloids  whence  they  came.  This  electric  charge  in  its  turn  reverses 
the  coagulative  change  in  the  colloids  and  so  brings  conditions  back 
towards  the  original  state.  Burridge  has  shown  that  the  positive 
charge  renders  cardiac  colloids  incapable  of  combining  with  Ca  and 
can  decalcify  them  just  as  well  as  does  oxalate,  and  that  Na  ions 
may  be  a  factor  in  determining  a  finer  state  of  subdivision  of  these 
colloids.  Inhibition  resides  in  the  inability  of  the  colloids  of  the 
inhibited  tissue  to  combine  with  Ca. 

Burridge  found  two  modes  of  reaction  of  the  heart  when  exposed 
to  the  influence  of  drugs.  One  effect,  immediate  in  appearance  on 
application  and  in  disappearance  on  withdrawal,  he  ascribes  to  "  sur- 
face" phenomena  involving  the  muscular  colloids.  The  other  he 
calls  "  deep  "  changes  on  the  assumption  that  they  involve  changes 
in  aggregation  or  of  chemical  composition  in  the  same  colloidal  bodies 
in  which  changes  of  the  first  type  take  place. 

Calcium  has  a  surface  and  a  deep  action  on  the  heart.  Burridge 
measured  the  response  of  the  perfused  heart  to  solutions  containing 
different  concentrations  of  calcium  in  the  presence  of  different  sub- 
stances. He  found  that  digitalis  caused  the  heart  to  act  as  well  with 
a  weaker  solution  of  calcium,  as  it  did  with  a  stronger  solution  of 
calcium  in  the  absence  of  digitalis.  Barium  was  an  imperfect  sub- 
stitute for  calcium.  Adrenin  and  pituitary  extract  act  like  digitahs 
in  improving  Ca  utihzation.  Alcohol,  chloroform  and  ether  have  a 
two-fold  action,  (a)  A  depress'ng  or  surface  action  associated  with 
poor  utihzation  of  calcium;  (6)  a  favoring  or  deep  action  associated 
with  an  improved  utihzation  of  calcium. 

Strychnine  improves  Ca  utihzation  thus  diminishing  inhibition. 
Dibasic  potassium  phosphate  increased  the  result  from  a  given  per- 
centage concentration  of  Ca.  This  latter  is  evidently  an  adsorption 
phenomenon  as  it  is  maintained  by  adding  a  smaller  percentage  of  the 


298b  colloids  in  BIOLOGY  AND  MEDICINE 

dibasic  potassium  phosphate  to  the  perfusing  solution.  Burridge 
suggests  that  loss  of  phosphates  may  occasionally  be  a  factor  deter- 
mining cardiac  failure.  The  antagonism  between  chlorids  and  phos- 
phates is  evidently  of  importance  in  cardiac  weakness  or  nephritis 
with  salt  retention.  W.  Burridge,  Quarterly  Journal  of  Medicine, 
Vol.  9,  Nos.  33  and  36;  Vol.  10,  No.  39  for  bibhography.  See  also 
Nerves,  p.  352.     Tr.] 


CHAPTER  XVIII. 
BLOOD,   RESPIRATION,   CIRCULATION  AND   ITS  DISTURBANCES. 

Blood. 

(See  also  Chapter  XIV,  The  Distribution  of  Water  in  the  Organism.) 

The  blood  consists  of  a  fluid,  the  plasma,  and  the  formed  elements, 
the  blood  corpuscles. 

Plasma. 

In  a  short  time  after  the  blood  leaves  the  body  it  coagulates 
spontaneously.  It  separates  into  two  components;  one  a  yellowish 
fluid,  the  serum,  and  the  jelly-like  clot,  the  fibrin,  which  has  enmeshed 
the  blood  corpuscles  and  has  undergone  shrinking  during  coagulation. 
If  the  blood  is  beaten  or  shaken  with  a  rough  surface  (wood  or  steel 
shavings)  after  it  has  left  the  vein,  it  clots  at  once  and  the  fibrin 
separates  immediately  as  an  irreversible  fibrous  mass  which  may  be 
completely  cleansed  of  the  adherent  constituents  of  the  blood  by 
washing  it  with  water.  Recently  colloid-chemical  explanations  asso- 
ciated with  the  names  K.  Spiro  and  Ellinger,  Nolf,  Rettger, 
and  Hekma  have  received  more  support.  Fibrin  occurs  in  the  blood 
as  fluid  fibrinalbuminate  (fibrinogen)  which  is  normally  characterized 
by  being  coagulated  by  dilute  salt  solutions  and  serum,  so  that 
something  must  exist  in  the  blood  which  prevents  coagulation  in  the 
vessels.  We  possess  no  knowledge  as  to  what  this  "something"  is. 
[Howell  has  recently  separated  this  substance  antiprothrombin.  See 
Harvey  Lectures,  1916-1917  to  be  published.  Tr.]  Certain  aspects 
may  be  indicated  which  help  the  solution  of  the  problem.  Coagula- 
tion occurs  when  the  blood  leaves  the  closed  vascular  system,  and 
comes  into  contact  with  other  surfaces  which  it  moistens.  If  blood  is 
collected  in  oil  or  vaseline,  it  remains  fluid  many  hours  and  may  even 
be  beaten  with  a  thoroughly  greased  glass  rod  without  clotting. 
Shed  blood  may  be  centrifuged  in  paraffined  vessels  and  a  plasma 
may  be  thus  obtained  which  remains  fluid,  provided  the  suggested 
precautions  are  employed.  Such  plasma  clots  when  a  glass  rod  which 
it  moistens  is  introduced.  It  is  not  known  whether  or  not  the  inter- 
face blood/gas  plays  any  part  in  clotting. 

H.  IscovESCO  is  of  the  opinion  that  the  electric  charge  of  the 
vessel  wall  plays  an  important  part  in  the  coagulation  of  fibrin;   in 

299 


300  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

life  it  differs  from  what  it  is  in  death  and  pathological  conditions. 
Blood  does  not  clot  in  a  paraffined  vessel  because  the  paraffin  is  nega- 
tively^ charged.  [The  spontaneous  and  immediate  development  of 
vasoconstrictor  substances  in  shed  blood  have  made  necessary  the 
development  of  special  technique  "  caval  pockets  "  for  the  study  of 
the  influence  of  the  nerves  on  the  adrenal  glands.  J.  M,  Rogoff, 
Journ.  Lab.  and  Chn.  Medicine,  Vol.  Ill,  No.  4,  Jan.,  1918,  p.  209 
et  seq.  Tr.]  He  arrives  at  this  view,  because  he  regards  clotted  fibrin 
as  a  complex  which  results  from  the  combination  of  all  the  electro- 
positive globulins  of  the  blood  with  some  of  the  negative  ones.  He 
assumes  that  plasma  contains  two  kinds  of  globulins,  one  of  which 
coagulates  at  72°  and  the  other  at  55°. 

At  any  rate  it  follows  from  what  has  been  said  that  the  determina- 
tive factor  in  coagulation  is  a  surface  tension  phenomenon  which 
offers  a  profitable  field  for  more  thorough  study.  It  must  not  be 
forgotten  that  the  blood  moistens  the  vessel  walls,  the  intima,  so 
that  it  is  not  the  mere  moistening  which  is  important.  Injuries  to 
the  intima  may  bring  about  local  blood  coagulation  {thrombosis). 
If  such  thrombi  are  dislodged  into  the  vessels  the  resulting  phenomena 
are  called  embolism.  It  is  well  known  that  the  chief  danger  of  any 
extensive  surgical  operation  is  the  development  of  such  emboli. 
Air  emboH  are  especially  dangerous;  they  may  occur  from  injuries 
to  the  veins,  or  air  may  be  injected  during  intravenous  infusions. 
Sometimes  in  air  embolism,  coagulation  may  occur  at  the  interface 
blood/air  as  I  have  been  informed  in  a  personal  letter  from  Geheimrat 
Prof.  Quincke,  the  clinician. 

The  serum  is  a  solution  of  proteins  in  salt  solution.  We  must  at 
present  assume  that  these  proteins  are  different,  not  only  for  every 
animal,  but  even  for  every  race.  They  consist  chiefly  of  serum 
albumin  and  serum,  globulin,  which  were  described  in  Chapter  X  (see 
also  IscovESCO*^).  It  is  quite  possible  to  remove  a  considerable 
portion  of  the  proteins  from  the  blood  without  immediate  destruc- 
tion of  the  organism;  as  is  well  known,  common  salt  infusions  are 
employed  in  severe  hemorrhages,  i.e.,  the  blood  that  is  lost  is  re- 
placed by  a  0.85  per  cent  salt  solution.  It  would  be  impossible  to 
keep  an  organism  alive  permanently  without  serum.  Aside  from 
the  fact  that  nourishment  would  cease,  the  serum  plays  a  most  im- 
portant role  as  "buffer"  in  order  that  the  acid  and  alkaH  content  of 
the  organism  may  be  kept  at  a  uniform  level;  it  is  of  little  impor- 
tance, however,  in  maintaining  the  level  of  the  water  content. 

^  French  authors  frequently  use  a  terminology  the  reverse  of  this;  they  call 
whatever  migrates  to  the  anode  electropositive,  and  vice  versa.  I  have  trans- 
lated their  mode  of  expression  so  as  to  conform  with  ours. 


BLOOD,  RESPIRATION,  CIRCULATION  AND  DISTURBANCES     301 

The  organism  is  carefully  provided  witli  mechanisms  to  maintain 
the  neutral  state.  [This  is  not  in  accordance  with  present  views. 
In  American  cHnical  literature  this  is  at  present  usually  expressed 
+  + 

H7,  Ph7  or  Ph7  =  neutral.     In  normal   blood  ps  =  7.4,   while  in 

advanced  acid  intoxication,  ^^7.2.  Tr.]  Every  abnormal  excess 
of  H  or  OH  ions  influences  the  condition  of  swelling  in  the  tissues 
and  may  thus  give  rise  to  grave  disturbances.  Although  0.37  X  10~^ 
represents  the  normal  H-ion  concentration  of  the  blood,  1.00  X  10~'^  nR 
indicates  an  advanced  acid  intoxication.  1.00  X  10"^  nH  damages 
the  red  blood  corpuscles  according  to  L.  Michaelis  and  D.  Tak- 
ABASHi  just  as  do  traces  of  NaOH.  We  know  from  the  inves- 
tigations of  H.  J.  Hamburger  and  Hekma*  that  the  functional 
activity  of  the  leucocytes  depends  on  the  normal  concentration  of 
H  and  OH  ions.  To  maintain  this  condition,  nature  has  provided 
a  double  safeguard  and  surrounded  neutralization  with  a  double 
hne  of  defense.  The  outer  wall  is  the  serum  salts,  which  are  so 
skillfully  combined  that  the  concentration  of  the  H  or  OH  ions  is 
unchanged  by  the  moderate  addition  of  acids  or  bases.  This  prop- 
erty of  the  serum  salts  is  of  great  importance  in  the  metabolism, 
formation  and  removal  of  CO2,  in  the  formation  of  lactic  acid,  of 
ammonia,  etc. ;  it  is  even  to  a  certain  extent  increased  by  the  artifi- 
cial introduction  of  acids  and  bases.  Circumstances  may  arise 
when  these  outworks  are  overcome  and  the  inner  Hne  trenches  have 
to  bear  the  defense. 

Severe  poisoning  with  acids  or  alkalis  are  dangerous  not  only 
because  of  the  burns  they  cause,  but  especially  because  of  the  danger 
of  disturbing  the  balance  between  the  H  and  OH  ions.  We  must 
especially  consider  the  acid  intoxication  in  certain  diseases,  as  in  the 
fever  of  many  of  the  infectious  diseases  and  in  diabetes,  associated 
with  the  over-production  of  oxybutyric  acid.  Under  these  circum- 
stances the  inner  though  weaker  line  of  defense  must  be  the  proteins, 
which,  because  of  their  amphoteric  character,  are  able  to  bind  acids 
as  well  as  bases.  It  is  not  a  matter  of  little  consequence  when  the 
proteins  have  to  be  invoked  for  neutralization.  We  have  seen 
from  pages  152  and  153  that  the  addition  of  alkali  or  acid  increases 
the  internal  friction  of  albumin;  that  albumin  ions  have  a  much 
higher  viscosity  than  the  albumin  molecule;  and  further  that  in 
media  that  are  not  neutral  the  blood  corpuscles  swell.  Therefore, 
under  circumstances  where  there  is  a  higher  ionization  of  serum 
albumin  and  parallel  with  it  a  swelling  of  the  formed  elements,  we 
may  expect  that  the  blood  vnW  show  a  higher  viscosity  and  that 
greater  demands  will  be  made  on  the  organ  of  circulation,  the  heart 
(see  p.  310  et  seq.). 


302 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


There  are  no  exhaustive  investigations  on  the  influence  of  salts 
upon  the  internal  friction  of  proteins  in  normal  serum.  Yet  it 
seems  possible  to  conclude  from  the  figures  of  Wo.  Pauli  and 
H.  Handovsky  that  the  friction  in  serum  in  the  presence  of  salts  is 
approximately  the  same  as  in  a  salt-free  albumin  solution;  however, 
it  must  not  be  forgotten  that  the  solutions  investigated  possessed  a 
much  smaller  amount  of  albumin  than  does  serum. 


Albumin  +  NaCl. 

Internal  friction. 

Albumin  +  (NH4)2S04 

Internal  friction. 

Normal. 

0.00 
0.05 
0.1 
0.5 

1.0783 
1.0592 
1.0681 
1 . 1064 

Normal. 
0.00 
0.05 
0.1 
0.5 

1.0783 
1.0582 
1.0725 
1 . 1020 

We  see  that  when  the  solution  of  salt  is  very  dilute,  the  internal 
friction  may  even  sink,  but  that  between  0.1  and  0.5  n  which  is  in 
the  neighborhood  of  the  physiological  concentration  (physiological 
salt  solution  0.85  per  cent  NaCl  =  0.14  normal),  the  original  inter- 
nal friction  of  the  salt-free  albumin  solution  is  again  reached  and 
exceeded, 

A  matter  formerly  very  much  discussed  was  whether  some  electro- 
lytes and  nonelectrolytes,  especially  sugar,  chlorin,  phosphate, 
sodium  and  calcium,  exist  in  the  blood  free  or  fixed.  In  the  light  of 
the  facts  this  does  not  seem  to  be  a  correct  statement  of  the  problem. 
It  is  more  important  to  determine  what  percentage  of  the  ions 
involved  are  diffusible,  B,  P.  Rona  and  György  by  ultrafiltration  of 
C02-sera  have  demonstrated  that  10  to  15  per  cent  of  the  Na  in  some 
sera  was  not  diffusible.  P.  Rona  and  D.  Takahashi  *  determined  that 
25  to  35  per  cent  of  the  calcium  in  the  serum  is  not  diffusible  and 
probably  exists  as  a  calcium  protein  compound.  The  CI  ion,  on  the 
contrary,  was  found  to  be  completely  diffusible. 

We  know  from  pages  147  and  161  that  the  solubility  of  salts  in 
solutions  of  hydrophile  colloids  is  very  different  from  what  it  is 
in  pure  water.  As  a  matter  of  fact,  the  solubility  of  easily  soluble 
electrolytes  is  somewhat  diminished  and  that  of  the  difficultly 
soluble  ones  is  markedly  increased.  If  we  make  a  solution  of  salts 
of  the  same  strength  and  proportion  per  liter  as  occurs  in  natural 
serum  (see  H.  M.  Adler  *) 

KCl 0.40 

CaCl2  +  6  aq : 0. 62 

MgCl2  +  6aq 0.37 

NaCl 5.90 

NaH,P04  +  1  aq 0.236 

NaHCOs 3.51 


BLOOD,  RESPIRATION,  CIRCULATION  AND  DISTURBANCES     303 

a  precipitation  takes  place  immediately,  the  precipitate  consisting  of  a 
mixture  of  calcium  carbonate  and  phosphate.  It  is  only  because  of 
the  presence  of  the  serum  colloids  that  this  precipitation  does  not 
occur.  Accordingly,  the  serum  colloids  serve  the  purpose  of  keeping 
soluble  substances  in  solution  and  of  releasing  them  at  the  proper 
time.  This  phenomenon  is  of  great  physiological  importance  in  the 
formation  of  bone  (see  p.  268),  and  it  is  of  great  pathological  signifi- 
cance in  gouty  deposits  (see  p.  274). 

The  surface  tension  of  serum  has  been  frequently  studied  in  recent 
years.  The  incentive  was  afforded  by  the  observation  of  M.  Ascoli 
and  G.  Izar  that  the  surface  tension  of  immune  serum  was  depressed 
when  it  was  united  with  its  specific  antigen  (see  meiostagmin  reaction). 
Morgan  and  Woodward  recorded  especially  exact  determinations. 
They  found  that  the  surface  tension  of  healthy  men  did  not  vary 
much,  on  the  average  (from  44.3  to  46.4),  but  that  the  diet  might 
cause  marked  variation.  The  surface  tension  is  especially  high  in 
some  patients,  especially  nephritics  (reaching  to  51.4).  Mammalian 
serum  does  not  differ  much  from  human  serum. 

Lymph. 

We  can  think  of  the  lymph  as  a  filtrate  derived  from  the  blood. 
It  does  not  by  any  means  have  a  composition  identical  with  blood 
plasma;  on  the  contrary,  we  find  in  it  many  metabolic  products  which 
have  entered  it  by  diffusion  from  the  cells  it  bathes.  It  is  generally 
assumed  that  the  blood  pressure  affords  the  increased  pressure 
necessary  for  filtration;  but  I  wish  to  call  attention  to  the  fact  that  it 
may  possibly  be  the  pulsation  which  is  of  prime  importance  in  this 
instance,  just  as  I  have  established  for  the  glomerular  filtration  in 
the  kidney  (p.  332). 

The  Blood  Corpuscles. 

The  red  blood  corpuscles  or  erythrocjrtes  present  under  the  mi- 
croscope the  well-knovm  round  or  elliptical  shape  with  a  thickened 
rim,  Hke  a  biconcave  circular  or  elliptical  plate.  In  reality,  according 
to  the  investigations  of  F.  Weidenreich,  they  seem  to  approach 
the  shape  of  a  spinning  top  (a  cone  with  a  convex  base) ;  so  that  under 
the  microscope  their  appearance  is  distorted.  When  they  leave 
the  blood  vessels  we  frequently  encounter  erythrocytes  in  rouleau. 
ScHWYZER  attributes  this  to  the  fact  that  the  normal  OH  charge  is 
disturbed  by  the  glass  shde.  In  the  blood  vessels,  however,  the  suni- 
lar  and  equal  charge  of  the  vessel  wall  opposes  the  corpuscles  and  they 
never  form  rouleau.  In  view  of  what  follows,  we  should  recall  that 
on  swelling,  they  swell  up  symmetrically  and  have  the  shape  of  a  pea, 


304  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

and  on  shrinking  they  take  the  shape  of  a  thorn  apple,  and  have 
characteristic,  pointed  outgrowths. 

Their  composition  varies  somewhat  according  to  the  species;  a 
beef's  red  blood  corpuscle,  besides  salts,  contains  (according  to  E. 
Abderhalden  *^) 

Per  mile. 

Water 591 . 6 

Hemoglobin 316.7 

Albumin. 64.2 

Cholesterin 3.4 

Lecithin 3.7 

In  an  electric  field  they  migrate  to  the  anode  (R.  Höber),  and 
though  as  a  matter  of  fact,  the  electric  charge  varies  according  to  the 
species  (Kozawa),  yet  H.  Iscovesco*^  considers  that  only  the  cap- 
sules and  the  stroma  are  electronegative,  because  intact  blood  cor- 
puscles and  stroma  are  precipitated  by  electropositive  iron  hydroxid 
sol.  The  content  on  the  contrary  is  electronegative,  since  corpuscles 
dissolved  in  water  are  precipitated  by  arsenic  sulphid  hydrosol. 

The  investigations  on  the  lowering  of  the  freezing  point  induced 
by  the  contents  of  red  blood  corpuscles,  etc.,  are  exceptionally 
numerous.  These  data  will  never  lose  their  value.  All  conclusions 
concerning  the  osmotic  pressure  of  the  respective  solutions  require  a 
revision  in  accordance  with  the  investigation  of  W.  Biltz  and  A. 
VON  Vegesack  (see  p.  46),  who  have  shown  that  the  presence  of 
colloids  greatly  influences  the  osmotic  pressure  of  crystalloids  (see 
H.  J,  Hamburger  and  F.  Bubanovic  *) .  It  is  established  by  the 
investigation  of  R.  Höber  that  er3rthrocytes  have  a  high  internal 
conductivity;  this  indicates  that  a  considerable  fraction  of  the  salts 
they  contain  are  freely  dissolved  and  do  not  occur  in  any  organic 
combination. 

As  numerous  as  have  been  the  studies  of  erythrocytes,  just  so 
divergent  are  the  ideas  concerning  their  structure.  Hitherto  it  has 
been  difficult  to  reconcile  the  demands  of  physical  chemistry  with 
their  other  properties.  By  alternate  freezing  and  thawing  again, 
hemolysis,  i.e.,  exit  of  coloring  matter  may  be  induced;  this  be- 
tokens a  capsule  which  may  be  burst  by  a  purely  mechanical 
effort  exerted  on  the  blood  corpuscle  in  its  entirety,  thus  giving  exit 
to  the  dissolved  hemoglobin.  This  capsule  must  for  the  most  part 
consist  of  fatty  colloidal  particles  (lecithin,  Cholesterin,  or  both), 
since  it  may  be  removed  by  ether  and  other  fat  solvents,  and  may 
even  be  melted  at  a  temperature  of  from  60°  to  65°  C,  thus  allow- 
ing the  hemoglobin  to  escape.  It  must  certainly  contain  lecithin 
since  pure  Cholesterin  cannot  be  moistened  by  water  and  is  totally 
impermeable  to  it.     Unfortunately,  the  properties  of  mixtures  of 


BLOOD,  RESPIRATION,  CIRCULATION  AND  DISTURBANCES     805 

lecithin  and  Cholesterin'  in  relation  to  water  and  salt  solution  are 
entirely  uninvestigated.  Investigation  of  the  sweUing  capacities  of 
such  mixtures  would  greatly  advance  our  knowledge  of  plasma 
membranes. 

It  is  evide^jt  that  the  lipoids  can  form  only  very  thin  surface  pel- 
licles if  we  consider  how  small,  in  view  of  the  analysis  on  page  304, 
is  the  amount  of  Cholesterin  and  lecithin  they  contain. 

A  question  remains:  Is  the  membrane  of  uniform  composition 
and  consistence?  Against  this  view  is  the  fact  that  on  shrink- 
ing, thorn-apple  forms  appear.  This  shape  might  also  occur  if 
there  were  an  irmer  framework,  the  interstices  of  which  are  filled 
at  the  periphery  with  compressible  elements.  Such  a  theory  best 
fits  in  with  physico-chemical  observations.  The  peUicle  of  red  blood 
corpuscles  is  permeable  for  water,  fat-soluble  substances  and  to  a 
certain  extent  for  cations  and  anions  of  the  salts  occurring  in  the 
body. 

The  existence  of  a  fatty  pellicle  does  not  eliminate  the  probability 
that  the  blood  corpuscles  may  also  form  a  fatty  layer  at  an  injured 
point,  which  impedes  the  exit  of  hemoglobin  (see  p.  243).  Gruns* 
discovered  that  red  blood  corpuscles  could  be  cut  up  without  the 
subsequent  exit  of  hemoglobin,  and  E.  Albrecht  *  showed  that  blood 
corpuscles  divided  by  crushing  or  by  gentle  heating,  to  a  certain 
extent  resumed  their  spherical  form  by  reason  of  surface  tension,  and 
retained  their  coloring  matter.  It  must  be  borne  in  mind  that  red 
blood  corpuscles  contaui  only  591  parts  water  to  316  parts  hemo- 
globin. If  the  hemoglobin  were  to  appropriate  all  the  water,  which 
is  certainly  not  the  case,  we  should  obtain  only  a  very  viscous  mass,i 
which  would  require  a  considerable  time  to  swell  or  undergo  other 
changes,  thus  giving  an  opportunity  for  the  formation  of  a  surface 
membrane  (see  p.  35). 

Based  on  my  own  hitherto  unpublished  studies  on  hemolysis  and 
on  those  of  others  accessible  to  me,  I  have  formulated  the  following 
hypothesis  for  the  structure  of  red  blood  cells:  they  possess  a  sponge- 
like framework  consisting  of  a  fibrinous  mass,  the  stroma  (to  what 
extent  lipoids  are  involved  in  this  framework  is  an  open  question 
entirely  immaterial  to  our  discussion).  The  sponge  is  entirely 
soaked  with  a  salt-containing  albumin  solution,  chiefly  hemoglobin, 
and  has  a  very  thin  hpoid  pellicle  as  a  capsule.  On  the  basis  of 
such  a  structure  all  the  known  properties  of  red  blood  cells  find  an 
unstrained  explanation.  They  would  swell  and  burst  in  hypotonic 
solution  and  shrink  in  hypertonic  solution.     Such  salt  solutions  as 

1  It  is  easy  to  convince  oneself  of  this  by  observing  a  pure  suspension  of 
blood  corpuscles  becomes  hemolyzed  by  a  drop  of  arachnolysin. 


306  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

dehydrate  (shrink)  the  stroma  without  causing  coagulation  of  the 
hemoglobin  hemolyze  red  blood  cells.  In  fact,  concentrated  sodium 
chlorid  solution  causes  hemolysis.  The  blood  coloring  matter  is 
pressed  out  just  as  in  the  case  of  the  salts  ^  of  heavy  metals  mentioned 
later.  By  carefully  cutting,  crushing  or  warming,  the  blood  corpus- 
cles do  not  lose  their  coloring  matter,  provided  sufficient  time  is 
allowed  for  the  lipoid  membrane  to  close  before  the  coloring  matter 
diffuses  out.  If  the  lipoid  membrane  be  destroyed  by  v/arming  or 
solvents  (ether,  saponin,  etc.),  hemolysis  occurs.  The  salts  of  the 
heavy  metals  which  coagulate  albumin  harden  red  blood  corpuscles 
also,  and  if  red  blood  corpuscles  are  placed  in  such  weak  solutions  of 
heavy  metals  that  no  coagulation  of  the  dissolved  albumin  occurs, 
there  are  two  possibihties :  the  salt  of  the  heavy  metal  causes  no 
shrinking  of  the  stroma  (e.g.,  CuClo)  and  the  blood  corpuscles  are  left 
unchanged;  or  the  heavy  metal  causes  shrinking  of  the  stroma  {e.g., 
HgCl2  and  many  others)  so  that  hemolysis  occurs,  for  the  blood  color- 
ing is  pressed  out  as  water  is  from  a  sponge. ^  The  spongy  frame- 
work with  its  largest  outgrowths  reaches  to  the  surface  of  the  blood 
corpuscles,  so  that  there  is  at  the  surface  if  there  is  no  stress  a  mosaic  of 
lipoids  and  albumins  which  can  assume,  in  hypertonic  neutral  salt  solu- 

1  With  a  few  special  exceptions,  it  is  entirely  immaterial  to  our  point  of  view 
whether  we  regard  behavior  towards  hypotonic  and  hypertonic  solutions  as  the 
result  of  osmotic  pressure  or  as  the  result  of  swelUng  and  shrinkage  of  the  corpus- 
cular coUoids. 

Martin  H.  Fischer  *  has  developed  an  entirely  revolutionary  conception  of 
the  constitution  of  the  red  blood  corpuscles  and  the  phenomenon  of  hemolysis. 
He  starts  out  with  the  idea  that  hemolysis  may  occur  as  two  phenomena:  (o) 
with  swelling  of  the  blood  corpuscles  (in  water,  acids,  alkalis,  etc.);  (b)  with- 
out swelUng  (alcohol,  saponin,  hemolysins,  etc.).  On  this  account  M.  H.  Fischer 
regards  the  increase  of  volume  and  the  exit  of  the  hemoglobin  as  two  phenomena 
which  frequently  run  parallel  and  yet  have  nothing  to  do  with  each  other.  He 
assumes  that  the  proteins  (not  the  hemoglobin)  swell  under  the  influence  of 
water,  acids,  alkahs  and  hypotonic  salt  solution.  The  hemoglobin,  however, 
which  he  considers  to  be  an  hydrophobe  colloid,  he  regards  as  being  adsorbed  by 
the  remaining  protein  constituents  of  the  blood  corpuscles.  To  demonstrate 
his  conception  M.  H.  Fischer  stained  fibrin  with  carmine  and  observed  a  loss  of 
color  with  acids,  alkahs,  hypotonic  salt  solutions,  urea,  etc.,  just  as  in  the  case 
of  hemolysis.  In  this  way  M.  H.  Fischer  injects  a  new  point  of  view  into  the 
discussion,  since  he  replaces  the  influence  of  osmotic  pressure  by  the  force  of 
sweUing  and  the  salts  by  the  colloids,  yet  many  of  his  points  seem  untenable  to 
me.  If  we  consider  that  a  blood  corpuscle  contains  almost  five  times  as  much 
hemoglobin  as  other  proteins,  we  must  cease  talking  of  an  adsorption  of  the 
hemoglobin  by  the  albumin;  adsorption  is  a  reversible  process,  a  term  applicable 
to  only  very  few  hemolytic  phenomena.  I  cannot  subscribe  to  the  view,  that 
hemoglobin  is  an  hydrophobe  (suspension  colloid).  This,  however,  is  not  vital 
to  the  main  question. 

2  From  experiments  still  unpublished. 


BLOOD,  RESPIRATION,  CIRCULATION  AND  DISTURBANCES     307 

tion,  a  thorn  apple  shape.  On  the  basis  of  this  hypothesis,  whenever 
we  exert  an  influence  on  red  blood  corpuscles,  we  must  consider  the 
effect  upon  each  of  the  three  colloidal  constituents  (stroma,  hemo- 
globin solution  and  lipoids)  as  well  as  its  distribution  among  them; 
from  this  the  behavior  of  the  blood  corpuscles  may  be  deduced 
(hemolysis,  swelling,  shrinking,  hardening,  etc.). 

There  is  an  extensive  literature  on  the  changes  in  the  volume  of 
erythrocytes  in  neutral  isotonic  salt  solutions;  ^  I  shall  mention  only 
the  names  of  Gürber,  H.  J.  Hamburger,  S.  G.  Hedin,  R.  Höber, 
H.  KoEPPE  and  M.  Oker-Blom.  In  these  studies,  the  blood  corpuscles 
were  usually  regarded  as  vesicles  with  a  more  or  less  permeable  mem- 
brane filled  with  a  solution  of  electrolytes.  This  conception  does  not 
permit  a  general  satisfactory  explanation  of  all  the  observations  that 
have  been  made.  There  are  already  evidences  of  a  revision  which 
shall  ascribe  due  influence  to  the  colloidal  character  of  the  corpuscular 
constituents.  R.  Höber  *''  immersed  blood  corpuscles  in  neutral  salt 
solution,  which  possessed  the  same  osmotic  pressure,  but  in  relation  to 
the  blood  corpuscles,  were  somewhat  hypotonic,  so  that  hemoglobin 
gradually  escaped.  This  escape  took  place  more  or  less  slowly  in 
accordance  with  the  salt  employed  and  in  the  following  order: 

SO4  <  CK  Br,  NO3  <  I 

Li,  Na  <  Cs,  Rb  <  K. 

This  is  the  recognized  lyotropic  arrangement  for  colloidal  pre- 
cipitation, or  what  seems  more  likely  to  me,  for  swelling  and  shrink- 
ing (see  M.  MicuLiciCH  *).  An  investigation  by  Eisenberg  contains 
much  important  data  which  ought  to  yield  valuable  conclusions,  in 
connection  with  the  theory  outlined  above.  [Note.  J.  Tait  has 
just  published  a  paper  —  Capillary  Phenomena  observed  in  Blood 
Cells:  Thigmocytes  Phagocytosis,  Ameboid  Movement,  Differential 
Adhesiveness  of  Corpuscles,  Emigration  of  Leucocytes.  Quarterly 
Journal  of  Experimental  Physiology,  Vol.  XII,  No.  1.     Tr.] 

Leucocytes  have  a  special  significance  which  we  have  considered 
in  Chapter  XVII. 

As  has  been  frequently  emphasized,  the  normal  organism  estab- 
lishes a  dynamic  balatice  in  the  swelling  of  the  organ  colloids  (see 
p.  217  et  seq.).  The  tissues,  the  blood  plasma,  the  blood  corpuscles 
possess  a  certain  swelling  range,  which  is  specific  for  each  tissue. 
If  certain  component  colloid  groups  suffer  disturbances  in  this 
respect,  in  order  to  restore  equilibrium  all  the  other  components 
modify  their  state  of  swelling.     This  may  occur  in  severe  diarrhoeas 

1  Numerous  investigations  have  been  performed  to  determine  the  behavior 
of  blood  corpuscles  towards  neutral  salt  solutions.  From  them  it  may  be  de- 
duced that  iso-osmoiic  solutions  are  not  necessarily  isotonic  to  blood  corpuscles. 


308 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


(cholera),  which  result  in  dehydration  of  the  entire  organism.  This 
is  combated  by  an  injection  of  physiological  salt  solution  into  the 
blood  vessels.  If,  on  account  of  abnormal  conditions,  there  is  an 
increased  swelHng  of  tissue  (edema,  exudates),  the  balance  may  be  re- 
stored by  withdrawing  water  from  the  blood  (by  sweating,  diuretics 
or  cathartics).  [Reference  should  be  made  to  the  influence  exerted 
by  increasing  the  colloids  of  the  blood  either  by  transfusion  of  blood 
or  by  injection  of  colloidal  substances.     Tr.] 

Respiration  (Gas  Exchange). 

The  supply  of  oxygen  to  the  cells  is  probably  the  most  important 
condition  for  the  life  of  the  organism,  whether  animal  or  plant .  For  the 
latter,  quantitative  estimations  are  not  as  convenient,  and  on  this  ac- 
count they  have  been  less  studied  than  in  animals,  especially  mammals. 

In  the  case  of  higher  animals,  the  provision  of  oxygen  is  assigned 
to  the  red  blood  corpuscles  which  take  up  oxygen  in  the  lungs  or  gills, 
transport  it  to  the  places  where  it  is  needed,  and  return  laden  with 
CO2.  Ability  to  take  up  and  relinquish  oxygen  or  carbon  dioxid  is  a 
characteristic  of  hemoglohm.  Formerly,  the  combination  of  oxygen  or 
carbon  dioxid  and  hemoglobin  was  considered  to  be  a  purely  chemical 
one.  In  favor  of  this  view  is  the  fact  that  it  is  possible  to  crystallize 
both  hemoglobin  and  also  oxygen-laden  oxyhemoglobin.  The  union 
must  be  a  very  loose  one,  since  it  is  possible  to  remove  with  an  ex- 
haust pump  almost  all  the  oxygen  from  an  oxyhemoglobin  solution  or 
even  from  oxyhemoglobin  crystals,  so  that  the  absorption  of  oxygen 
and  carbon  dioxid  follows  the  gas  pressure.  It  would  be  natural  to 
think  of  a  solution  of  0  or  CO2  in  the  hemoglobin,  but  quantitative 
investigations  show  in  contradiction  that  the  absorption  of  0  or  CO2 
is  not  proportional  to  the  outer  gas  pressure  as  would  occur  for  the 
solution  of  a  gas  in  a  fluid  in  accordance  with  Henry's  law.  It  has 
been  found  on  the  contrary,  that  with  low  gas  pressures  relatively 
much  O  or  CO2  is  taken  up,  but  that  with  higher  pressures  the  amount 
diminishes;  the  following  table  of  A.  Loewy*^  shows  this: 


Oxygen  tension  in 
mm. 

Oxygen  saturation  in  per 
cent  with  a  CO2  ten- 
sion of  5  mm. 

5 

11 

10 

28.5 

15 

51 

20 

67.5 

30 

82 

40 

89 

50 

92.5 

80 

98 

100 

99 

150 

100 

BLOOD,  RESPIRATION,  CIRCULATION  AND  DISTURBANCES     309 

Since  Bonder's  time  it  has  been  assumed,  therefore,  that  a  disso- 
ciation occurs  much  as  in  the  case  of  calcium  carbonate  which  is 
dissociated  into  CaO  and  CO2  at  high  temperatures;  the  degree  of 
dissociation  is  dependent  upon  the  CO2  pressure.  Chr.  Bohr  es- 
pecially followed  up  this  idea  and  on  the  basis  of  rather  complicated 
premises  reached  an  approximate  agreement  between  theoretical 
and  experimental  values.  H,  W.  Fischer  and  E.  Brieger  think 
that  iron  exists  in  hemoglobin  "  protected  "  by  the  organic  complex. 
In  alkaline  solution  it  occurs  as  a  ferrate,  i.e.,  a  superoxid  (analogous 
to  manganate),  but  in  a  solution  made  acid  by  CO2  it  is  unstable  and 
parts  with  oxygen.  Another  explanation  offered  by  the  assumption 
of  Wo.  OsTWALD  *^  is  that  the  taking  up  and  release  of  0  or  CO2 
by  hemoglobin  and  blood  is  an  adsorption.  He  compares  the  process 
to  the  absorption  of  gases  by  charcoal,  spongy  platinum,  etc.  In 
both  cases  there  exists  a  reversible  balance,  in  both  cases  one  gas 
may  be  replaced  by  the  other  (0  by  CO2  and  vice  versa).  The  curves 
obtained  in  the  adsorption  of  0  or  CO2  by  hemoglobin  or  blood  in 
the  presence  of  increasing  gas  pressures  correspond  to  the  recog- 
nized adsorption  curves.  On  comparing  the  differences  between 
theory  and  experiment  in  the  case  of  Bohr's  dissociation  formula, 
on  the  one  hand,  and  Wo.  Ostwald's  adsorption  formula,  on  the 
other,  it  is  evident  that  for  the  latter  they  are  much  smaller. 

Hitherto  we  have  spoken  of  the  adsorption  of  O  and  CO2  by 
hemoglobin,  blood  corpuscles  and  blood  as  identical,  but  in  reality  the 
phenomena  in  the  blood  are  very  comphcated. 

The  absorption  and  the  release  of  oxygen  by  hemoglobin  may  be 
considered  a  relatively  simple  phenomenon.  But  even  in  the  case  of 
blood  corpuscles  and  blood,  observation  and  calculation  do  not  agree 
so  well.  Wo.  OsTWALD  says  (loc.  cit.,  p.  296),  "These  variations  are 
characterized  by  the  fact  that  in  the  case  of  low  oxygen  pressures 
(somewhat  below  25  mm.  Hg)  the  observed  oxygen  absorption  is  con- 
siderably less  than  would  be  expected  in  accordance  with  the  adsorp- 
tion formula. "  In  my  opinion  this  variation  is  sufficiently  explained  if 
we  consider  the  lipoids  of  blood  corpuscles;  0  is  more  soluble  in  them 
than  in  water  and  there  is  no  objection  to  the  assumption  that  the 
distribution  in  the  lipoids  with  changing  gas  pressure  follows  Henry's 
law.  Under  these  circumstances  the  curve  of  oxygen  absorption  in 
the  blood  must  waver  between  that  of  the  adsorption  curve  and  the 
straight  line  of  distribution  in  accordance  with  Henry's  law.  By 
observation  of  the  curve  elaborated  by  Wo.  Ostwald  (in  accordance 
with  A.  Loewy),  I  find  this  view  substantiated  (loc.  cit,  p.  297). 

The  absorption  and  release  of  CO2  by  the  blood  and  blood  corpus- 
cles is  further  comphcated  by  the  blood  salts.    By  their  presence  the 


310  COLLOIDS  IN  BIOLOGY  AßD  MEDJCINE 

serum  is  able  to  take  up  more  CO2  than  the  blood  corpuscles,  but  it  is 
impossible  at  present  to  formulate  in  detail  the  steps  in  the  absorption 
and  release  of  CO2  by  the  blood.  The  adsorption  theory  of  Wo. 
OsTWALD  naturally  does  not  help  us  over  these  difficulties,  but  under 
the  compHcated  conditions  mentioned  it  formally  estabhshes  thfe 
adsorption  character  of  the  absorption  and  release  of  CO2  in  the  blood. 
If  we  view  teleologically  the  manner  of  gas  exchange,  we  recog- 
nize that  adsorption  serves  this  purpose  best.  Excess  of  oxygen 
(up  to  100  per  cent)  in  the  respired  air  has  no  effect  either  in  the 
0  used  or  on  the  general  metabohsm;  when  oxygen  is  deficient  small 
quantities  are  taken  up  with  avidity  and  tenaciously  held.  The  in- 
halation of  oxygen  recently  recommended  for  clinical  use  may  be 
explained  only  by  considering  the  plasma.'^  From  a  mixture  rich  in 
oxygen,  hemoglobin  will  take  up  only  a  given  maximum  quantity, 
but  the  ability  of  the  plasma  to  take  up  oxygen  follows  the  gas 
pressure.  Finally,  it  must  be  recalled  that  with  higher  pressure  the 
fipoids  of  the  blood  corpuscles  take  up  more  oxygen.  [No  discussion 
of  the  gas  exchange  in  the  blood  would  be  complete  without  mention 
of  the  work  of  Barcroft,  "  Respiratory  Function  of  the  Blood," 
Cambridge,  1914,  and  of  Henderson,  and  of  Donald  van  Slyke, 
who  have  supplied  methods  and  data  of  great  value.     Tr.] 

The  Circulation  and  Its  Disturbances. 

A  normal  circulation  can  exist  only  when  the  internal  friction  of 
the  blood,  the  viscosity,  remains  within  normal  limits.  This  varies 
considerably,  and  in  man,  measured  for  uncoagulated  blood,  is  from 
4.05  to  6.8  (water  =  1).  In  cardiac  patients,  besides  normal  values, 
values  from  above  14  to  23.8  have  been  found.  Muscular  exercise, 
heat  aiid  chemical  stimuli  influence  the  viscosity  (H.  A.  Deter- 
MANN*).  Salivation,  lack  of  water,  perspiration  increase,  whereas 
ingestion  of  fluids  or  nourishment  as  well  as  increased  frequency 
of  respiration  lower  the  viscosity  (M.  Scheitlin*).  According  to 
H.  Blunschly  the  viscosity  of  the  blood  falls  with  every  intake  of 
nourishment,  reaching  a  minimum  after  the  midday  meal,  and  then 
rises  with  fluctuations.  The  differences  in  the  same  person  on  one 
day  were  11.8  per  cent;  yet  the  figures  vary  much  on  different  days. 
Moderate  muscular  work  according  to  H.  A.  Determann  lowers 
the  viscosity  of  the  blood,  hard  work  raises  it  as  do  alcohol  and 
coffee. 

The  viscosity  of  the  blood  may  be  influenced  by  certain  substances; 
according  to  W.  Scheitlin,*  a  gelatin  injection  of  0.15  per  cent  of  the 
1  Naturally  this  does  hold  in  the  case  of  CO  poisoning. 


BLOOD,  RESPIRATION,  CIRCULATION  AND  DISTURBANCES     311 

blood  volume  increases  the  viscosity  temporarily  15  per  cent;  are- 
colin  (0.1  grm.,  subcutaneously)  up  to  36  per  cent;  cutaneous  appli- 
cation of  Spiritus  sinapis,  up  to  12  per  cent;  a  phlebotomy  (up  to 
12  per  cent)  and  the  action  of  other  derivatives,  for  a  few  hours 
lower  the  viscosity  (these  results  were  obtained  on  horses).  Changes 
in  viscosity  were  observed  by  W.  Scheitlin  in  various  diseases 
(principally  of  horses) ;  and  by  W.  Frei  *  on  dying  horses.  In  dis- 
eases of  the  lungs  and  pleura  associated  with  fever,  especially  high 
values  were  found,  and  in  anemias  they  were  especially  low  (as  low 
as  2.3).  The  viscosity  usually  reaches  its  highest  point  with  the 
crisis  and  then  falls. 

All  these  observations  furnish  valuable  material  for  future  knowl- 
edge of  the  relationship  between  the  viscosity  of  the  blood  and 
pathology.  Already  it  may  be  said  that  the  viscosity  of  the  blood 
has  a  certain  prognostic  value. 

The  viscosity  of  the  blood  is  conditioned  by  the  internal  friction 
of  the  plasma  and  by  the  blood  cells.  We  shall  see  later  that  the 
total  amount  of  the  former  plays  a  far  less  important  part  than 
the  latter,  and  that,  as  a  matter  of  fact,  an  important  part  in  the 
changes  in  viscosity  of  the  plasma  must  also  be  ascribed  to  the 
interface  tissue/plasma. 

Marked  changes  in  the  viscosity  of  the  plasma  may  also  be  indu  ced 
by  the  inclusion  of  some  foreign  substances.  P.  Adam  found  that  it 
is  lowered  markedly  by  iodids,  and  to  a  less  extent  by  bromids. 
It  is  possible  that  some  of  the  therapeutic  effects  from  the  adminis- 
tration of  potassium  iodid  (especially  in  arteriosclerosis)  may  be 
attributed  to  the  diminution  of  the  internal  friction.  Investigations 
on  man  have  not  as  yet  given  uniform  results  (P.  Adam,  H.  A. 
Determann,*  O  Müller  and  R.  Inada). 

The  concentrations  of  urea  possible  in  the  organism  can  have  no 
influence  on  the  internal  friction  of  the  plasma  (as  far  as  I  can  gather 
from  G.  MoRuzzi's  *  figures). 

To  the  extent  that  the  data  furnished  by  Wo.  Pauli  and  H. 
Handovsky  warrant  it,  an  increased  ionization  of  the  serum  albumin 
must  also  result  in  an  increase  of  viscosity.  Such  an  increased 
ionization  of  albumin  may  be  brought  about  by  an  increase  in  con- 
centration of  H  or  OH  ions.  An  increase  of  the  latter  is  impossible, 
as  we  shall  see.  On  the  other  hand,  the  following  tables  of  R.  Höber 
show  that  an  increase  of  CO2  may  produce  an  increase  of  the  con- 
centration of  H  ions  in  the  blood. 

R.  HÖBER*-  measured  the  concentration  of  H  ions  in  the  blood 
upon  the  addition  of  carbonic  acid  mixed  with  hydrogen.  I  have 
reproduced  only  the  percentage  of  CO2. 


312 


COLLOIDS  IN  BIOLOGY  AND   MEDICINE 


Volume  per  cent  CO2. 

Concentration  of 
H  ions  X  10' 

3.18 
4.15 
6.51 

9.19 
15.50 
29.05 

57.86 

0.37 
0.49 
0.79 
0.89 
0.94 
2.37 
2.98 

Within  the  physiological  limit  of  3  to  6  volume  per  cent  CO2,  the 
concentration  of  H  ions  in  the  blood  varied  only  from  about  0.35  to 
0.75  X  10"^  When  the  content  of  CO2  is  greater,  we  shall  see  that 
the  H  ion  concentration  may  rise.  According  to  H.  Loewy,*^  the 
normal  CO2  content  in  the  alveoli  on  closure  of  the  air  passages  may 
increase  from  5  per  cent  to  13.4  per  cent,  which  produces  approx- 
imately 50  per  cent  increase  in  the  concentration  of  H  ions. 

A.  SziLi  *  injected  rabbits  and  dogs  intravenously  with  hydro- 
chloric acid  and  found  shortly  before  death  an  H  ion  concentration 
of  9  X  10~^;  in  diabetic  coma  H.  Benedict*  observed  a  ps value  of 
1.5  X  lO-l 

This  increase  in  the  H  ions  is  doubtless  associated  with  an  in- 
creased viscosity  of  the  plasma.  We  shall  see  that  this  is  vanishingly 
small,  hardly  measurable  in  proportion  to  the  increase  of  the  vis- 
cosity of  the  blood  which  an  equal  concentration  of  H  ions  produces 
through  a  swelling  of  the  red  hlood  corpuscles. 

The  viscosity  of  the  total  plasma  does  not  teach  us  anything  about 
its  viscosity  at  the  interface  between  tissue  and  blood.  The  friction 
at  this  surface  is  absolutely  determinative  for  the  circulation  of  the 
blood. 

What  are  the  conditions  at  these  surfaces?  The  blood  is  neutral, 
the  tissues  are  acid.  In  the  cells  there  is  an  oxidation  which  passes 
bj^  way  of  the  most  diverse  fatty  acids  to  carbonic  acid.  The  fatty 
acids  involved  are  without  exception  stronger  acids  than  carbonic 
acid.  Accordingly,  at  the  interface  tissue/blood,  an  ionization  of 
the  albumin  must  occur  and  with  it  an  increase  of  the  friction.  The 
friction  must  be  greater  in  proportion  to  the  disturbances  of  the  oxidiz- 
ing processes  in  the  cells;  i.e.,  the  less  oxidation  to  CO2,  the  faintly  acid 
end  product,  the  more  acids  with  high  dissociation  constants  are 
formed.  The  friction  at  the  boundary,  i.e.,  the  albumin  ionization, 
becomes  great  also  when  the  blood  itself  is  saturated  with  CO2,  that  is, 
when  the  products  which  diffuse  into  the  blood  from  the  tissues  have 
less  alkali  to  combine  with  than  normally. 


BLOOD,  RESPIRATION,  CIRCULATION  AND  DISTURBANCES     313 

Let  us  test  the  correctness  of  this  view  with  the  facts  at  our  dis- 
posal.^ 

It  follows  from  the  evidence  to  be  given  (p.  315)  that  the  accumu- 
lation of  CO2  increases  the  viscosity  of  the  blood,  but  we  do  not  learn 
from  this  how  the  swelhng  of  the  blood  corpuscles  and  to  what  ex- 
tent the  H  ion  concentration  at  the  interface  tissue/blood  may  be 
involved.  Indirectly  we  obtain  a  certain  insight  from  the  studies 
of  acid  intoxications.  A.  Loewy  and  E.  Munzer*  determined  the 
abihty  of  the  blood  to  absorb  CO2  in  normal  animals  and  in  those 
poisoned  with  hydrochloric  acid,  with  the  following  results. 


Normal  Blood. 


CO2  tension. 

CO2  fixation. 

Per  cent. 

2.196 
3.290 

Volume  per  cent. 
28.43 
34.75 

Blood  of  an  Animal  Poisoned  with  Acid. 

CO2  tension. 

CO2  fixation. 

Per  cent. 
3.630 
6.143 
7.530 

Volume  per  cent. 

7.37 

17.88 

22.26 

Accordingly,  the  CO2  tension  must  be  greater  in  the  animal 
poisoned  with  acid  for  the  same  absorption  by  the  blood  to  occur 
as  in  a  normal  animal;  which  means  that  there  is  less  alkali  for  use 
in  combining  with  the  acid  products  diffusing  from  the  tissues  than 
in  the  normal  one. 

Similar  conditions  occur  when  there  is  an  abnormal  acid  produc- 
tion in  the  tissues;  namely,  after  great  muscular  exertion  with  over- 
production of  lactic  acid,  in  fever,  where,  measuring  the  alkalinity 
of  the  blood  (by  the  CO2  exhausted).  Kraus  found  it  reduced  to  one- 
half  or  one-third  of  the  normal,  as  in  typhoid,  erysipelas,  scarlatina 
or  continued  fever  of  tuberculosis,  in  starvation,  in  coma,  especially 
diabetic  coma  with  over  production  of  oxybutyric  acid,  and  frequently 
in  diabetes  meUitus.     Kraus  found  in  a  severe  case  instead  of  a  vol- 

1  It  is  only  possible  to  iodicate  here  that  the  difference  in  the  reaction  be- 
tween blood  and  tissue  necessitates  a  difference  in  potential  which  offers  resist- 
ance to  the  movements  of  the  blood.  It  is  greater  in  proportion  to  the  relative 
difference  in  the  concentration  of  H  ions.  Unfortunately,  we  lack  the  experi- 
mental basis  to  establish  my  assumption  mathematically. 


314  COLLOIDS   IN  BIOLOGY  AND  MEDICINE 

ume  per  cent  from  30  to  36  CO2  only  12.4  and  9.8,  and  O.  Minkowski 
once  found  only  3.3  per  cent.  [Mention  should  be  made  of  the  work 
of  JosLiN,  VAN  Slyke,  Marriot  and  Rowland  in  America.   Tr.] 

On  these  grounds  we  must  realize  that  in  all  respiratory  dis- 
turbances not  only  the  accumulation  of  CO2  in  the  blood,  but  also 
that  the  incomplete  oxidation  in  the  tissue  resulting  from  insufficient 
O,  leads  to  circulatory  disturbances. 

The  following  line  of  proof  seems  especially  interesting  to  me. 
We  know  that  in  obesity,  the  oxidizing  forces  in  the  tissues  are  re- 
duced, so  that  the  fat  is  no  longer  attacked,  and,  as  a  matter  of 
fact,  it  is  in  the  obese  especially  that  circulatory  disturbances  regu- 
larly appear. 

Clinicians  know  that  the  circulatory  disturbances  from  diminished 
alkalescence  in  such  cases  are  benefited  by  giving  large  doses  of 
alkalis. 

I  am  fully  convinced  that  the  facts  heretofore  mentioned  may 
be  explained  by  the  increased  viscosity  of  the  blood  due  to  the 
swelUng  of  the  blood  corpuscles,  and  also  that  we  have  no  experi- 
mental evidence  to  separate  the  two  phenomena.  My  sole  object 
is  to  introduce  into  the  organic  development  of  existing  ideas  a 
new  viewpoint  based  on  colloid-chemical  facts.  At  the  interface, 
tissue/hlood,  increased  H  ion  concentration  may  develop,  and  as  a  result 
of  the  ionization  of  the  albumin,  a  greater  friction  may  he  produced. 
The  increase  in  the  concentration  of  H  ions  may  be  produced  by 
diminution  in  the  alkalinity  of  the  blood  or  through  a  disturbance  of  the 
oxidizing  processes  in  the  tissues,  so  that  many  circulatory  disturb- 
ances may  be  viewed  as  errors  of  metabolism.  The  possibility  of  an 
increased  friction  as  the  result  of  a  change  in  the  difference  in  potential 
between  blood  and  tissue  has  thus  been  indicated. 

The  mere  disturbance  in  the  oxidation  processes  in  the  tissues  also 
causes  them  to  swell,  resulting  in  an  abstraction  of  water  from  the 
blood  and  a  consequent  increase  in  its  viscosity.  We  must  thus 
recognize  a  relationship  between  the  viscosity  of  the  blood  and  the 
questions  considered  in  the  chapter  on  Edema  (see  p.  223  et  seq.). 

The  influence  of  the  blood  cells  on  the  internal  friction  of  the  blood. 
As  was  shown  by  A.  Gürber,  the  red  blood  corpuscles  swell  when 
carbonic  acid  is  introduced.  H.  J.  Hamburger  and  von  Limbeck 
have  confirmed  and  thoroughly  analyzed  this  observation  in  the  case 
of  carbonic  acid  and  other  acids.  The  increase  in  the  volume  of  the 
red  blood  corpuscles,  which  is  15  per  cent  more  in  venous  than  in 
arterial  blood,  in  pathological  conditions,  e.g.,  in  asphyxiation,  may 
rise  to  30  per  cent. 

The  most  exhaustive  investigations  in  this  direction  were  under- 


BLOOD,  RESPIRATION,  CIRCULATION  AND  DISTURBANCES     315 

taken  by  A.  von  Koranyi  and  J.  Bence.  It  is  to  the  credit  of  A. 
VON  Koranyi  that  he  appUed  these  results  to  the  disturbances  of  the 
circulation.  We  have  to  thank  him  for  a  truly  illuminating  dis- 
cussion of  the  pathological  physiology  of  the  circulation  (A,  von 
Koranyi  and  0.  Richter,*  Vol.  II,  p.  51  ef  seq.). 

Finally,  we  may  conclude  that  increase  in  the  CO2  content  causes  a 
swelling  of  the  blood  corpuscles  which  may  be  considered  the  chief 
factor  in  the  increased  viscosity  of  the  blood.  By  introducing 
oxygen  the  process  is  reversed. 

Circulatory  disturbances  may  be  conditioned  by  failure  of  the 
motor,  the  heart,  by  changes  in  the  pipe  system,  the  arteries,  veins 
or  capillaries,  or  finally  by  increased  viscosity  of  the  blood.  Every 
change  in  the  internal  friction  of  the  blood  must  in  the  first  place 
have  an  effect  upon  the  heart,  and  in  the  second  place  upon  the 
tubular  system.  It  follows  then  that  a  deficient  cardiac  function 
may  primarily  produce  an  increased  viscosity  of  the  blood  (through 
insufficient  supply  of  oxygen),  or  an  increased  viscosity  of  the  blood, 
due  primarily  to  a  disturbance  in  tissue  metabolism,  may  secondarily 
result  in  a  disturbance  of  the  heart.  We  have  recognized  that  an 
accumulation  of  acids,  especially  an  accumulation  of  CO2,  may  be 
responsible  for  an  increase  in  the  viscosity  of  the  blood.  It  would 
indeed  be  a  grave  error  were  we  to  believe  that  the  course  of  events 
in  the  body,  during  circulatory  disturbances,  has  the  simple  formula 
we  have  given.  A  further  presentation  of  the  complicated  circum- 
stances and  the  therapeutic  influences  would  take  us  far  beyond  the 
limits  of  this  book,  although  most  of  the  phenomena  have  not  as  yet 
been  considered  from  the  colloid-chemical  standpoint.  The  increase 
in  the  number  of  red  t)lood  corpuscles,  the  increase  in  their  CI  con- 
tent, etc.,  the  entire  course  of  circulatory  decompensation  and  com- 
pensation are  questions  which  in  the  present  state  of  knowledge  are 
solved  better  by  the  practised  eye  of  clinicians  than  by  the  calcula- 
tions of  research. 

Secretion  and  Absorption. 

Water  and  food  enter  the  gastrointestinal  canal  in  normal  nutri- 
tion. The  food  is  changed  from  a  colloidal  to  a  crystalloidal  con- 
dition and  is  thus  able  to  pass  through  the  intestinal  membrane  with 
the  water  and  so  reach  the  interior  of  the  body  —  it  is  absorbed. 
Excess  of  water  and  useless  crystalloids  are  eliminated  by  the  glands 
(secretion)  _  If  we  accept  this  rough  sketch,  secretion  becomes,  as 
M.  H.  Fischer  has  defined  it,  the  mirror  image  of  absorption.  There 
are  organs  which  take  up  solutions  and  those  that  eliminate  them. 
We  know  from  previous  chapters  that  the  organism  strives  its  utmost 


316  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

to  maintain  its  normal  condition  of  swelling,  so  that  there  must  be 
phenomena  in  the  absorptive  organs  which  oppose  those  in  the 
secreting  glands.  M.  H.  Fischer  recognized  these,  both  in  the 
oxidation  processes  of  the  cells  and  in  the  circulation.  The  func- 
tionating cell  producing  acid,  like  venous  blood  rich  in  CO2,  has  an 
increased  swelling  capacity  —  it  absorbs  water;  the  resting  cell, 
being  well  supplied  with  oxygen  like  the  arterial  blood,  has  an  excess 
of  water  —  it  secretes.  Thus  we  see  that  in  the  fluctuating  supply  of 
oxygen  to  an  organ  are  furnished  the  conditions  for  the  occurrence 
of  absorption  and  secretion. 


CHAPTER  XIX. 
ABSORPTION. 

(See  also  p.  409  et  seq.,  section  on  Purgatives.) 

The  absorption  of  food  materials  or  alien  substances  is  accom- 
plished by  a  dissolving  current  which  originates  in  the  intestines  and 
flows  through  the  body.  There  it  parts  with  some  substances  and 
takes  up  others,  finally  excreting  through  the  various  glands,  but 
especially  the  kidneys,  such  crystalloids  as  have  become  superfluous. 

Substances  may  be  absorbed  from  other  places,  through  the  skin 
and  mucous  membranes,  from  peritoneum  and  pleura.  This  may 
occur  when  exudates  have  collected  and  have  been  absorbed,  or  when 
the  substances  have  been  injected.  In  some  animals,  as  in  the  frog, 
absorption  of  water  occurs  only  through  the  skin.  Subcutaneous, 
intramuscular  and  intravenous  injections  are  made  in  order  to  intro- 
duce substance  into  the  body  without  their  passage  through  the 
intestines.  The  absorption  of  substances  thus  introduced  into  the 
organism  is  termed  parenteral  absorption. 

Alimentary  Absorption. 

Besides  the  fluids  which  are  taken  with  food,  there  is  daily  poured 
into  the  intestines  of  adult  men  700  to  1000  c.c.  of  sahva,  600  to 
900  c.c.  of  bile,  600  to  800  c.c.  of  pancreatic  juice,  1000  to  2000  c.c.  of 
gastric  juice,  200  c.c.  of  succus  entericus;  in  all  3.1  to  4.9  liters. 
Inasmuch  as  in  healthy  men  hardly  400  to  500  c.c.  of  this  fluid  are 
evacuated  with  the  feces,  there  must  normally  occur  a  reabsorption 
of  2.7  to  4.5  liters.  To  this  must  be  added  1  to  1.5  liters  of  hquid  food, 
so  that  the  alimentary  tract  absorbs  about  6  liters  daily  —  quite  a 
considerable  task. 

With  the  water,  dissolved  substances  are  also  absorbed.  Inas- 
much as  the  intestinal  membrane  is  impassable  for  colloids  (with 
the  exception  of  very  finely  emulsified  fats),  it  is  the  function  of 
the  alimentary  tract  with  the  help  of  the  ferments  to  convert  the 
foodstuffs  into  an  easily  diffusible  condition. 

The  intestinal  tube  is  a  membrane,  which  separates  the  interior 
of  the  body  from  the  foods  introduced  and  the  digestive  juices.  It 
is  the  function  of  research  to  discover  what  forces  drive  these  solutions 

317 


318 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Of  crystalloids  through  the  intestinal  membrane.  For  many  years 
it  was  believed  that  osmotic  forces  were  involved.  This  conception 
proved  to  be  a  serious  obstacle  to  further  progress  since  it  diverted 
attention  in  a  direction  offering  no  prospect  of  results.  It  was  only 
when  swelling  and  shrinking  were  recognized  as  the  dominant  factors 
in  absorption  that  it  became  possible  to  understand  many  experi- 
mental data  which  previously  had  been  inexplicable. 

The  recognition  of  the  importance  of  swelling  for  absorption  dates 
from  F.  Hofmeister.  He  said,  ''  The  essentially  absorbing  appara- 
tus, the  intestinal  epithelium,  is  possessed  of  the  power  to  swell. "  ^ 


Water  and  Crystalloids. 

If  water  or  dilute  salt  solution  is  introduced  into  a  loop  of  in- 
testines, it  is  absorbed  more  or  less  rapidly.  If  the  intestine  be- 
haved like  a  parchment  membrane  and  the  absorption  of  the  fluids 
was  brought  about  by  the  osmotic  pressure  of  the  body  juices,  water 
would  be  most  rapidly  absorbed.  This  is,  however,  not  the  case. 
According  to  G.  O.  Gumilevskij,*  pure  water  is  less  rapidly  absorbed 
than  0.25  per  cent  NaCl  solution.  Accordingly,  the  intestine  behaves 
Uke  gelatin  which  swells  more  in  salt  solution  than  in  pure  water. 

The  matter  becomes  more  complicated  if  we  consider  quantita- 
tively the  absorption  of  water  and  salt  in  hypotonic  and  hypertonic  salt 
solutions.  We  shall  study  the  experimental  figures  of  M.  Heiden- 
hain, who  placed  sodium  chorid  solutions  of  various  concentrations 
in  loops  of  small  intestine  of  a  dog  and  permitted  them  to  remain 
there  for  15  minutes. 


Introduced. 

Recovered. 

Experiment. 

c.c. 

Per  cent 
NaCl. 

Total  quan- 
tity NaCl. 

c.c. 

Per  cent 

NaCl. 

Total  quan- 
tity NaCi, 

1 

2 
3 

4 

120 
120 
117 
120 

0.3 
0.5 
1.0 

1.46 

grams 

0.36 
0.6 
1.17 
1.75 

18 

35 

75 

109 

0.60 
0.66 
0.90 
1.20 

grams 

0.108 
0.23 
0.67 
1.31 

A  glance  suffices  to  show  that  this  experiment  cannot  be  explained 
by  the  osmotic  relations:  with  a  parchment  tube  having  slight  swell- 
ing capacity  surrounded  by  physiological  salt  solution,  the  amount 
of  water  recovered  would  have  to  be  more  than  117  and  120  c.c. 

^  I  would  venture  the  suggestion  that  many  poisons  which  are  supposed  to 
lose  their  "physiological  components"  by  absorption  through  the  intestines,  such 
as  NaFl,  osmic  acid,  etc.,  markedly  modify  the  swelling  capacity. 


ABSORPTION  319 

respectively  (instead  of  75  and  109  c.c.)  in  experiments  3  and  4. 
The  process  becomes  clear  inomediately  if  we  regard  absorption  as 
actually  a  swelling  phenomenon.  We  know  from  page  67  that  ac- 
cording to  F.  Hofmeister,  a  swollen  gelatin  absorbs  more  salt  than 
water  from  a  dilute  salt  solution,  and  that  in  the  presence  of  NaCl 
the  absorption  of  water  is  stronger  than  in  the  case  of  pure  water. 

Further  light  is  thrown  on  this  discovery  by  the  researches  of  M. 
Oker-Blom,*  who  showed  that  serum  takes  up  a  hypertonic  common 
salt  solution  more  readily  than  an  isotonic  one. 

From  the  table  on  page  318  we  see  that  a  concentrated  salt  solution 
which  causes  shrinking  is  but  slowly  absorbed. 

If  we  consider  the  absorption  of  other  salts,  we  find  that  those 
which  as  a  rule  promote  the  swelling  of  fibrin,  gelatin,  etc.,  also  as 
a  rule  promote  the  absorption  of  water  'm  the  intestines.  Therefore, 
there  is  a  further  parallelism  between  rapidity  of  diffusion  and  fur- 
therance of  swelling. 

From  the  researches  of  R.  Höber  *i  as  well  as  those  of  G.  B.  Wal- 
lace and  A.  R.  Cushny,*  we  obtain  the  following  series: 

Rate  of  diffusion ;!   HPO4,  SO4  <  Fl  <  NO3  <  I  <  Br  <  CI, 
Rate  of  absorption:   Fl  <  HPO4,  SO4  <  NO3  <  I  <  Br  <  CI, 
Rate  of  diffusion :  Mg  <  Ca  <  Ba  <  Na  <  K, 
Rate  of  absorption :  Ba  <  Mg,  Ca  <  Na,  K. 

From  this  series  we  see  that  there  is  in  general  a  parallehsm  between 
the  rates  of  diffusion  and  of  absorption. 

Fl  and  Ba  being  powerful  protoplasmic  poisons,  it  does  not  sur- 
prise us  that  they  form  an  exception  and  inhibit  absorption. 

Similar  relationships  between  the  rates  of  diffusion  and  of  absorp- 
tion were  determined  for  a  series  of  organic  salts  and  nonelectrolytes. 

It  may  be  said,  then,  that  slowly  diffusing  substances  are  slowly 
absorbed,  and  that  electrolytes  causing  shrinking  may  impede  not 
only  their  own  absorption  but  also  that  of  others,  and  that  substances 
rapidly  diffusible  and  favoring  swelling  act  contrariwise.  This  follows 
from  the  investigations  of  F.  Hofmeister  and  his  pupils  as  well  as 
those  of  H.  Bechhold  and  J.  Ziegler. 

The  importance  of  swelling  is  especially  evident  when  two  different 
substances  are  absorbed  simultaneously.  Let  us  take  for  example 
an  experiment  of  Katzenellenbogen.*  Simultaneously  with  so- 
dium chlorid,  there  were  introduced  glycocol  and  acetone,  which 
surely  do  not  favor  swelling,  but  the  latter  rather  the  reverse,  and 
urea  which  induces  swelling  to  a  high  degree. 

^  For  I,  Br  and  CI  there  have  been  substituted  the  dififusion  path  in  jellies 
instead  of  rapiditj  of  diffusion. 


320 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Introduced. 

Recovered. 

Experiment. 

c.c. 

c.c.  of  menstruum 
(average). 

Per  cent  NaCl. 

1 

2 
3 

Glycocol+0.45%NaCl 
Acetone  +0.45%  NaCl 
Urea         +0.45%  NaCl 

50 
50 
50 

36 

15.5 

17 

0.332 
0.631 
0.496 

From  this  we  see  that  sodium  chlorid  absorption  is  greatest  in 
the  presence  of  urea,  because,  in  spite  of  the  great  shrinkage  in  the 
quantity  of  fluid,  the  concentration  of  sodium  chlorid  is  but  slightly 
increased.  This  coincides  with  the  results  of  the  experiments  of 
H.  Bechhold  and  J.  Ziegler  *^  in  which  urea  in  the  main  favors 
diffusion  in  gelatin  and  in  jellies.  If  we  assign  to  acetone  properties 
similar  to  alcohol,  we  may  explain  the  surprisingly  rapid  absorption 
of  water  in  the  above  experiment,  because,  as  has  been  said  on  page 
70,  a  certain  proportion  of  alcohol  increases  the  ability  of  glue  to 
swell.  From  these  observations  we  gather  new  points  of  view  in 
regard  to  the  ready  absorption  of  protein  cleavage  products,  which  1 
shall  elaborate  on  page  411. 

We  may  make  still  further  deductions  from  the  above  observations, 
since  in  experiment  1  we  have  seen  that  the  NaCl  content  of  the 
blood  is  higher  (about  0.6  per  cent);  so  that  it  has  been  absorbed 
against  the  osmotic  pressure  of  NaCl  in  the  intestines.  This  is  ex- 
plained on  pages  318  and  319.  In  experiment  2,  the  contrary  is 
the  case:  the  osmotic  pressure  is  higher  than  in  the  blood;  appar- 
ently the  structures  capable  of  swelhng  absorb  less  NaCl  in  the 
presence  of  acetone. 

Nor  need  we  be  surprised  by  the  fact  discovered  by  Otto  Cohn- 
HEiM,  that  a  hypotonic  grape  sugar  solution  introduced  into  a  loop 
of  the  small  intestine  becomes  more  concentrated.  From  the  same 
experiments  we  know  that  grape  sugar,  which  of  itself  diffuses  slowly, 
decreases  the  permeability  of  the  swollen  jelly,  and  thus  blocks  its 
own  passage. 

Substances  which  themselves  possess  great  capacity  to  swell,  e.g., 
agar,  hinder  the  absorption  of  water  to  a  high  degree. 

It  still  remains  a  question,  how  the  intestine  maintains  its  ability 
to  take  up  water,  since  water  is  constantly  being  removed  from  the 
intestine  by  diffusion  or  swelling.  The  theory  proposed  by  Martin 
H.  Fischer  *  seems  to  offer  valuable  assistance. 

He  starts  with  the  idea  that  venous  blood  containing  carbonic 
acid  has  a  tendency  to  swell,  i.e.,  to  take  up  water;  arterial  blood,  on 


ABSORPTION  321 

the  other  hand,  has  a  tendency  to  shrink  or  become  dehydrated.  He 
showed  that  the  arteries  of  the  mesentery  spread  out  into  a  capillary 
network  which  lies  directly  under  the  intestinal  epithelium  and  empties 
into  the  portal  vein  as  blood  containing  much  carbonic  acid.  Ac- 
cording to  V.  Limbeck,  A.  Gürber  and  H.  J.  Hamburger  red  and 
white  blood  corpuscles  undergo  an  increase  in  volume  of  from  5  to  10 
per  cent  when  they  enter  the  venous  blood  from  the  arteries.  Ac- 
cording to  this,  one  liter  of  blood  which  passes  through  the  intestine 
would  take  up  17.5  c.c.  of  water  even  if  we  were  to  ascribe  the  ab- 
straction of  water  to  the  blood  corpuscles  alone.  From  this  fact, 
M.  H.  Fischer  assumes  that  venous  blood,  so  long  as  it  is  present 
as  such,  absorbs  water  from  the  intestinal  mucous  membrane.^ 

Though  the  facts  so  far  discovered  seem  so  clearly  to  explain  the 
processes  of  absorption,  we  must  not  overlook  the  fact  that  they  are 
the  result  of  experiments  which  have  nothing  in  common  with  natural 
taking  of  food,  etc.,  by  mouth.  On  this  account,  certain  objections 
have  been  raised.  The  living  intestinal  wall  is  not  a  closed  tube  but 
one  traversed  by  solutions,  so  that  it  is  therefore  a  question  whether 
many  phenomena  are  not  quite  different  in  the  living  animal.  The 
important  discovery  of  Loeper  *  must  be  mentioned :  if  salt  solutions 
of  a  given  concentration  are  given  by  mouth,  they  are  either  concen- 
trated or  diluted  so  that  they  reach  the  intestines  in  approximately  iso- 
tonic condition.  From  a  practical  standpoint,  all  those  conclusions 
must  be  ignored  which  are  based  on  the  presence  of  hypotonic  or  hyper- 
tonic salt  solutions.2     For  the  pharmacologic  deductions  see  page  411. 

Within  the  intestine  the  osmotic  pressure  must  vary  greatly  with 
the  enzymatic  cleavage  of  the  food,  and  it  is  impossible  to  under- 
stand why  at  low  osmotic  pressures,  salts  of  the  blood,  or  crystal- 
loid cleavage  products  of  albumin  might  not  diffuse  back  into  the 
intestine  from  the  interior  of  the  body.  Based  on  the  investiga- 
tions of  F.  Abderhalden,  it  is  almost  certain  that  colloidal  albumin 
is  reconstructed  from  its  cleavage  products  in  the  intestinal  wall. 
The  intestinal  wall  functionates  as  a  suction  pump  for  the  crystalloid 
cleavage  products  of  albumin  which  draws  a  stream  of  crystalloids 
from  the  intestinal  lumen  in  the  direction  of  the  interior  of  the  body. 
We  may  dismiss  from  consideration  the  absorption  of  albumins. 

1  In  my  opinion,  Fischer's  theory  offers  valuable  aid  in  explaining  nervous 
influences  upon  processes  of  secretion  and  resorption.  I  might  suggest  that  nervous 
diarrheas  may  be  attributed  to  increased  arterial  blood  supply  to  the  mesentery. 
It  is  quite  natural  to  regard  diarrheas  accompanying  inflammatory  processes  in 
the  intestines  as  a  consequence  of  the  increased  supply  of  arterial  blood. 

2  In  my  opinion  this  does  not  exclude  the  fact  that  when  introduced  into  the 
intestines,  the  intestinal  contents  are  usually  hypertonic  since  crystalloids  are 
uninterruotedly  formed  as  a  result  of  digestion  in  the  intestines. 


322  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

There  are  two  important  facts  established  by  H.  J.  Hamburger  *^ 
and  GiRARD.*  The  former  showed  that  it  was  possible  to  prepare 
membranes  that  were  more  permeable  from  one  side  than  from  the 
other.  As  a  result  of  this  experiment  of  H.  J.  Hamburger  we  may 
state  that  crystalloids  pass  from  the  lumen  of  the  intestine  toward  the 
periphery  and  yet  no  crystalloids  of  the  blood  may  diffuse  back  into 
the  lumen  of  the  intestine.  It  remains  an  open  question,  whether 
the  small  quantities  of  NaCl  found  in  the  intestine  in  absorption 
experiments  come  from  the  secretion  of  the  intestinal  glands,  or 
whether  the  semipermeability  of  the  intestinal  membrane  is  limited 
in  the  direction  of  the  lumen  of  the  intestine. 

Of  the  greatest  general  importance  are  the  investigations  of 
GiRARD.  The  exhaustive  theoretical  discussion  of  their  basis  would 
lead  us  too  far  afield.  The  following  experiment  is  suggested:  if  a 
salt  solution  is  suspended  in  a  pig's  bladder  in  water,  the  rate  of 
diffusion  depends  on  whether  the  salt  solution  is  neutral,  slightly 
alkaline  or  slightly  acid.  The  cause  of  this  is  found  in  the  fact  that 
the  membrane  is  the  seat  of  an  electromotive  force.  We  conclude 
from  this  that  by  changing  the  reaction  on  both  sides  of  the  intestinal 
membrane  the  rate  of  diffusion  may  be  changed  or  regulated;  so 
that,  e.g.,  with  an  alkaline  reaction  which  occurs  in  the  intestines, 
the  diffusion  is  very  strongly  increased. 

Hitherto  we  have  started  from  the  assumption  that  the  intestinal 
wall  is  of  uniform  texture.  This  is  not  the  case,  as  a  view  of  a  micro- 
scopic section  and  simple  consideration  shows;  the  intestine  is  com- 
posed of  cells  as  is  every  other  organ,  and  we  know  that  cells  are 
only  permeable  for  crystalloids  to  a  limited  extent.  Accordingly, 
there  must  exist  an  easier  way  for  the  absorption  of  crystalloids  than 
through  the  cells;  it  must  occur  intercellularly.  It  is  different  in  the 
case  of  lipoid-soluhle  substances;  they  are  absorbed  intracellularly, 
e.g.,  ethyl  alcohol  is  absorbed  much  more  rapidly  than  salt  solution. 
Numerous  experiments  of  R.  Höber  have  established  in  general  the 
correctness  of  this  view. 

The  colloid-chemical  study  of  intestinal  absorption  has  already 
yielded  results  for  pathology. 

E,  Mayerhofer  and  E.  Pribram,*  in  a  series  of  valuable  inves- 
tigations, have  tested  the  condition  of  swelling  and  the  ability  to 
swell  of  the  normal  and  the  pathologically  changed  intestine. 
They  found  that  the  normal  and  especially  the  acutely  inflamed 
intestine  were  much  swollen  and  the  latter  possessed  an  increased 
permeability  for  crystalloids.  The  chronically  inflamed  intestine, 
which  already  showed  a  connective  tissue  atrophy,  had  a  very  small 
swelling  capacity,  so  that  a  much  more  limited  passage  was  afforded 


ABSORPTION  323 

to  dissolved  substances.'  Outside  the  body,  excised  pieces  of  intes- 
tine, when  artificially  swollen,  showed  an  increased  permeability 
for  KCl,  NaCl  and  dextrose  such  as  is  possessed  by  acutely  in- 
flamed intestine.  The  reverse  occurred  on  partial  drying  (shrink- 
ing); a  diminished  permeability  of  the  pieces  of  intestine  v^as 
obtained  corresponding  to  chronic  enteritis.  By  means  of  dehy- 
drating substances  (alcohol,  tannin)  it  was  possible  to  remove  the 
great  difference  in  permeability  between  the  acutely  swollen  and 
the  chronically  swollen  intestine.  Sugar  (dextrose),  especially, 
markedly  increases  the  permeability  of  the  intestinal  wall. 

Thus,  the  clinical  discoveries  of  H.  Finkelstein,  concerning  in- 
toxication with  sugar-rich  mixtures  in  children,  find  colloid-chemical 
confirmation.  [An  interesting  colloid-chemical  appHcation  is  the 
treatment  of  diarrheas,  especially  in  children,  by  adjusting  the  diet 
with  a  view  to  the  formation  of  insoluble  calcium  soaps  in  the  intes- 
tine. It  has  also  been  pointed  out  by  Bosworth  et  al.,  Am.  Jour. 
Diseases  of  Children,  Vol.  XV,  No.  6,  p.  397  et  seq.,  that  the  forma- 
tion of  Ca  soaps  may  interfere  with  the  absorption  of  fats  and  account 
for  certain  types  of  malnutrition  and  milk  intolerance.     Tr.] 

We  must  refrain  here  from  a  more  exhaustive  investigation  of 
absorption  by  the  stomach.  The  number  of  experimental  observa- 
tions is  still  not  large  and  from  a  colloid-chemical  standpoint  they 
do  not  assist. 

It  is  established,  e.g.,  that  the  stomach  absorbs  no  water  (von 
Mering)  ;  on  the  contrary  dissolved  substances  (salts,  carbohydrates 
and  albumins)  above  a  "certain  level  of  concentration"  are  absorbed 
(for  bibliography  see  H.  Strauss  *^  and  W.  Roth  and  H.  Strauss  *). 

Parenteral  Absorption. 

Like  those  concerning  intestinal  absorption,  the  studies  of  paren- 
teral absorption  have  been  based  almost  exclusively  on  the  laws  of 
osmotic  pressure  in  relation  to  the  permeabihty  of  membranes  (see 
U.  Friedemann*^). 

The  countless  difficulties  were  only  successfully  met  with  the 
entrance  of  the  colloid-chemical  views  of  swelling  and  shrinking,  for 
which  the  researches  of  Martin  H.  Fischer  *  paved  the  way. 

Researches  on  the  absorption  from  serous  cavities  have  been 
most  numerous.  As  regards  the  absorption  of  exudates,  the  ques- 
tion should  be,  under  what  circumstances  do  exudates  form?  Nor- 
mally the  organism  allows  no  fluid  to  collect  in  the  serous  cavities. 
We  must,  therefore,  assume  a  change  in  the  permeability  of  the 
living  membrane  or  a  shrinking  of  the  surrounding  tissue."^ 
1  See  also  M.  H.  Fischer,  "Nephritis." 


324  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

On  the  basis  of  M.  H.  Fischer's  investigations,  the  latter  view 
must  be  regarded  as  correct.  If  a  dilute  salt  solution  is  placed  in 
the  peritoneal  cavity  of  a  normal  living  or  a  dead  guinea  pig,  it  will 
soon  be  absorbed.  More  concentrated  salt  solutions  as  well  as 
solutions  of  the  salts  which  cause  loss  of  water  in  the  case  of  fibrin, 
gelatin,  etc.  (e.g.,  sodium  sulphate,  citrate  and  tartrate)  are  either 
not  absorbed  at  all  or  even  increase  their  volume  inasmuch  as  fluid 
enters  the  peritoneal  cavity  from  the  body.  If  albumin  solution 
is  injected  into  the  peritoneal  cavity,  very  little  will  be  absorbed 
within  an  hour.  The  tissues  about  the  peritoneal  cavity  behave  in 
this  case  just  as  does  every  dead  colloid  that  is  capable  of  swelling, 
and  there  is  no  difference  between  intestine  and  peritoneal  cavity. 
The  results  are  not  so  uniform  when  glycerin  or  sugar  solution  are 
injected  into  the  peritoneal  cavity.  Though  these  have  a  very  slight 
influence  on  the  swelling  of  fibrin,  they  cause  an  entrance  of  fluid 
into  the  peritoneal  cavity.  Evidently  they  cause,  as  they  do  in  the 
intestine,  an  irritation  (see  p.  323). 

The  investigations  of  this  question  are  not  completed  as  the  re- 
searches of  L.  AsHER  *  have  shown. 

A  great  number  of  marine  animals  have  a  body  surface  which  is 
permeable  for  water  though  not  for  salts.  Corals,  echinoderms, 
holothurians,  etc.,  swell  in  dilute,  and  shrink  in  concentrated  sea 
water.  [A  practical  application  is  the  "ripening  "  of  oysters."  Tr.] 
The  same  fact  obtains  for  many  fresh-water  animals,  especially  am- 
phibia, worms  and  snails.  Frogs  never  take  water  through  their 
mouths,  they  "drink  through  their  skins."  In  the  case  of  these  fresh 
water  animals  there  occurs  no  swelling  during  life  for  the  kidney 
excretes  the  excess  of  water.  The  skin  is  also  permeable  for  lipoid- 
soluble  substances.  The  skin  of  warm-blooded  anitnals  is  different 
from  that  of  cold-blooded  animals  in  that  it  is  almost  impermeable 
for  water  and  allows  only  lipoid-soluble  substances  to  pass  through 
(W.  FiLEHNE  *  and  A.  Schwenkenbecher*). 

From  what  has  been  said,  it  is  evident  that  a  whole  group  of 
factors  such  as  rate  of  diffusion,  swelling,  osmotic  pressure,  play  a 
part  in  absorption  —  and  we  shall  observe  the  same  for  secretion  — - 
and  that  there  are  many  gaps  in  the  experimental  data  which  sup- 
port these  views. 

The  undoubtedly  active  role  of  adsorption  has  not  been  touched 
upon;  and  it  is  possible  that  the  negative  phase  of  the  blood  pul- 
sations may  have  certain  importance  through  its  suction.  Some 
phenomena  regarded  as  filtrations,  for  which  the  pressure  in  the  in- 
testines seems  somewhat  too  slight,  may  be  explained  by  a  pulsating 
loss  of  swelling  (dehydration).     The  alcohol  question  received  a  new 


ABSORPTION  325 

light  from  colloid  studies.  We  saw  on  page  70,  that  gelatin  swelled 
more  in  weak  alcohol  solutions,  and  we  know,  on  the  other  hand, 
that  strong  alcohol  causes  shrinking.  Thus  we  may  possibly  ex- 
plain the  favorable  action  that  small  doses  of  alcohol  have  on  the 
absorption  of  nourishment.  Some  day,  chronic  alcoholism  may  pos- 
sibly receive  a  physico-chemical  explanation  from  the  change  in  the 
condition  of  the  body  colloids,  whether  albuminous  or  lipoid. 

[W.  BuRRiDGE  has  offered  what  seems  an  adequate  physico-chemical 
explanation  of  chronic  alcoholism.  Quarterly  Jour,  of  Medicine, 
Vol.  X,  No.  39.  In  the  alcohohc  who  "takes  a  httle,  often,"  the 
adrenin-like  action  of  alcohol  predominates,  and  there  is  an  improved 
utilization  of  Ca.  To  accommodate  for  this  the  Ca  tension  of  the 
blood  is  diminished  so  that  when  deprived  of  his  drug  the  circulation 
of  the  chronic  alcoholic  is  like  one  using  a  perfusion  solution  containing 
an  inadequate  supply  of  Ca.  This  accounts  for  the  acute  circulatory 
symptoms  upon  the  withdrawal  and  possibly  for  the  tremor.  It  also 
accounts  for  the  increased  anesthetic  risk,  if  the  accustomed  Ca 
tension  in  the  blood  is  below  that  which  is  adequate  to  maintain  the 
circulation  in  the  presence  of  a  percentage  concentration  of  anes- 
thetic usual  for  anesthesia.  A  similar  colloid-chemical  explanation 
based  on  this  work  of  W.  Burridge  may  be  offered  for  other  drug 
addictions.     Tr.l 


CHAPTER  XX. 

SECRETION  AND   EXCRETION. 

(See  also  Chapter  XIII,  The  Enzymes.) 

The  Glands. 

The  lymph  and  the  blood  current  discharge  into  the  glands, 
where  they  are  modified  in  a  specific  manner.  The  result  of  this 
selective  action  is  the  secretion  which  pours  from  the  ducts  of  the 
various  glands. 

The  cause  of  secretion  is  one  of  the  most  debated  of  physiolog- 
ical questions.  None  of  the  blood  colloids  are  found  in  the  secre- 
tions, but  the  crystalloids,  which  are  also  contained  in  the  blood, 
occur  in  abundance.  The  freezing  point  depression  gives  us  an  idea 
of  the  total  crystalloid  content.  This  shows  that  the  crystalloid 
content  (expressed  in  osmotic  pressure)  of  the  saliva  is  always  lower 
than  that  of  the  blood;  the  osmotic  pressure  of  gastric  juice  and  bile 
may  equal  that  of  the  blood;  milk  has  approximately  the  same 
osmotic  pressure  as  blood  and  fluctuates  with  it  in  this  respect. 
Sweat  and  especially  urine,  on  the  contrary,  may  either  have  a  lower 
osmotic  pressure  than  the  blood  or  exceed  it.  If  secretion  were  only 
an  ultrafiltration  of  the  blood  through  the  gland  filters,  it  would  be 
incomprehensible  how  an  excretion  could  occur  with  a  lower  osmotic 
pressure  than  the  blood,  or  with  one  that  is  higher,  as  sweat  and 
urine.  The  conditions  are  especially  complicated  by  reason  of  the 
fact  that  the  relative  crystalloid  content  of  various  secretions  may 
be  absolutely  different  from  what  they  are  in  the  blood  plasma. 
Thus,  for  instance,  milk  contains  from  4  to  5  per  cent  milk  sugar, 
whereas  the  blood  shows  only  from  0.08  to  0.12  per  cent  grape  sugar; 
urine  contains  disproportionately  more  urea  than  other  salts,  whereas 
the  amount  of  urea  in  the  blood  is  entirely  insignificant.  We  thus 
find,  besides  producing  specific  substances  (bile,  pepsin,  ptyalin,  etc.), 
that  the  individual  glands  have  a  specific  activity  in  relation  to  the 
lymph  crystalloids,  which  some  biologists  even  to-day  prefer  to  assign 
to  the  inexplicable  physiological  functions,  and  thus  to  remove  it 
from  physical  and  chemical  study.  As  yet  colloid  chemistry  has  no 
better  explanation  to  offer.  However,  it  seems  probable  to  me  that 
we  shall,  with  its  help,  come  to  a  solution  of  these  questions. 

326 


SECRETION  AND  EXCRETION  327 

We  may  assume  that  various  colloid-tissue  elements  differ  in  their 
ability  to  absorb  different  crystalloids  which  they  remove  from  solu- 
tion, just  as  fibers  remove  dyes,  and  that  thus  a  different  composition 
is  given  to  the  ultrafiltrate.  There  is  ample  reason  to  believe  that 
certain  crystalloids  (e.g.,  urea,  nitrates,  etc.)  open  the  paths  through 
the  colloid  membrane,  while  others  {e.g.,  sulphates)  close  them  just  as 
was  shown  in  their  diffusion  experiments  with  jellies  by  H.  Bechhold 
and  J.  ZiEGLER  (see  p.  55).  Side  by  side  with  this  occur  the  specific 
chemical  activities  of  the  gland  elements  which  give  them  their  par- 
ticular character,  the  formation  of  ptyalin  in  the  salivary  glands, 
and  of  bile  in  the  liver,  etc. 

E.  Pribram  *^  has  formulated  a  theory  according  to  which  there 
first  occurs  a  coagulation  of  nutritive  material  (granule  formation) 
in  the  gland  cells,  which  is  followed  by  swelling,  i.e.,  secretion.^ 

With  the  above  systematization  of  glandular  function,  we  may  ob- 
tain an  explanation  of  a  number  of  processes,  and  we  may  learn  how 
to  study  the  remainder.  We  shall,  therefore,  regard  every  secretion 
as  an  ultrafiltrate  whose  composition  is  changed  by  reabsorption  and 
specific  adsorption,  and  to  which,  in  the  case  of  most  glands  ^  (e.g., 
salivary,  pancreas,  liver,  etc.),  specified  chemical  products  of  glandu- 
lar activity  are  added.  In  what  order  the  change  by  adsorption 
and  ultrafiltration  occurs  remains  for  the  present  an  open  question. 

The  presence  of  free  water  is  an  essential  condition  for  the  ultra- 
filtration of  the  blood.  This  applies  only  to  the  glands.  As  was 
mentioned  in  more  detail  elsewhere,  we  may  assume  with  Maütin  H. 
Fischer  that  venous  blood,  rich  in  CO2,  abstracts  water,  whereas 
arterial  blood  can  release  it.  We  now  know  that  all  glands  are 
plentifully  supplied  with  arterial  blood  so  that  the  free  water  neces- 
sary for  ultrafiltration  is  supplied.  The  influence  of  the  pulsations 
of  blood  pressure,  noted  by  H.  Bechhold,  is  considered  on  page  332. 

^  Many  authors  distinguish  between  secretion  and  excretion  (urine,  sweat, 
etc.);  whereas  the  former  contain  colloidal  ingredients,  they  are  more  or  less 
completely  absent  from  the  latter.  Since  there  is  no  essential  difference  we  shall 
not  strictly  enforce  this  classification. 

2  It  cannot  be  denied,  that  the  consideration  of  the  digestive  glands  from  this 
viewpoint  offers  very  considerable  difficulties,  since  their  function  is  controlled 
to  a  very  great  extent  by  nervous  influences.  The  glands  which  are  not  directly 
under  the  control  of  the  nervous  system,  e.g.,  the  kidnej^s,  give  clearer  pictures. 
Since  we  know  that  there  is  an  excess  of  water  in  arterial  blood  and  that  venous 
blood  fixes  water,  the  nervous  control  of  secretion  becomes  a  working  hj'pothesis 
or  at  least  the  question  becomes  shifted,  and  we  must  investigate  the  eflfect  of 
nervous  influences  on  the  supply  of  blood  to  the  glands. 


328  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  Saliva. 

Under  normal  circumstances,  700  to  1000  c.c.  of  saliva  are  secreted 
daily.  The  amount  of  salivary  secretion  is  largely  influenced  by 
the  amount  of  water  contained  in  the  body.  If  large  amounts  of 
water  are  taken,  the  salivary  secretion  is  plentiful,  whereas,  with 
diarrhea,  profuse  sweating  and  fever,  it  diminishes  greatly  (dry 
mouth). 

Of  all  the  secretions,  the  saliva  has  on  the  average  the  lowest 
osmotic  pressure;  its  freezing-point  depression  is  from  0.11°  to  0.27°  in 
man,  in  contrast  to  blood  with  from  0.58°  to  0.60°.  If  the  excretion  of 
saliva  increases,  there  is  an  increase  in  the  crystalloids  contained  and 
to  such  an  extent  that  when  the  secretion  is  greatest  it  almost  reaches 
that  of  blood  plasma;  it  then  contains  0.58  per  cent  NaCl.  The 
fact  that  with  increased  salivation  the  carbonates  are  proportionately 
increased  in  the  saliva,  strongly  supports  in  my  opinion  the  role  of 
ultrafiltration  of  the  blood  plasma  in  the  secretion  of  the  saliva.  If 
the  NaCl  content  of  the  blood  is  increased,  that  of  the  saliva  also 
increases,  and  conversely  (J.  Novy,  J.  N.  Langley  and  Fletcher). 
If  potassium  iodid  or  Hthium  citrate  are  introduced  into  the  cir- 
culation (J.  N.  Langley  and  Fletcher),  iodin  or  lithium  may  be 
demonstrated  in  the  saHva  immediately;  whereas,  after  introducing 
grape  sugar  and  potassium  ferrocyanid  they  do  not  appear  in  the 
sahva  at  all  or  only  after  a  long  time.  All  these  facts  support  the 
view  that  the  chief  process  is  an  ultrafiltration.  The  last-mentioned 
experiments,  especially,  show  that  potassium  iodid  and  lithium  citrate, 
which  diffuse  rapidly  and  open  paths,  appear  in  the  sahva  promptly, 
whereas  grape  sugar  and  potassium  ferrocyanid,  as  a  result  of  their 
slowness  in  diffusion,  penetrate  the  filter  membrane  slowly,  so  that 
meanwhile  they  may  be  excreted  in  other  ways.  For  the  two  remain- 
ing elements  m  the  function  of  the  sahvary  glands,  the  change  in  the 
composition  of  the  ultrafUtrate  and  the  addition  of  the  colloid  con- 
stituents, we  have  as  yet  no  experimental  data. 

The  Bronchial  Glands.^ 

An  analysis  of  the  individual  functions  of  the  secretion  of  the 
bronchial  mucous  membrane  has  proved  hitherto  entirely  impossible. 
It  is  an  indication  of  ultrafiltration  that  with  an  increase  in  the 

^  Martin  H.  Fisher  with  justice  questions  why  the  secretion  of  superfluous 
water  commences  in  the  kidneys  instead  of  in  the  lungs,  since  by  the  change  of 
the  venous  blood  rich  in  carbonic  acid  into  arterial  blood,  water  is  actually 
Hberated.  He  thinks  that  water  is  not  secreted  in  the  lungs  because  of  the  im- 
permeabihty  of  the  membrane,  or  because  there  is  insufficient  time. 


SECRETION  AND  EXCRETION  329 

secretion,  there  is  an  increase  of  the  alkali  carbonates  in  the  mucus, 
which  like  all  alkaline  substances  fluidify  the  mucus  colloids, 
especially  the  mucin.  The  utilization  of  potassium  iodid  as  an  ex- 
pectorant may  be  similarly  interpreted;  possibly  it  also  favors  ultra- 
filtration in  the  sense  of  H.  Bechhold  and  J.  Ziegler.  Moreover, 
R.  HÖBER  upon  applying  iodin  found  an  increase  in  the  ciliary  move- 
ments in  the  ciliated  epithelium  of  frogs;  this  increased  activity 
may  assist  in  the  discharge  of  mucus. 

Gastric  Juice. 

The  daily  excretions  of  the  stomach  amount  from  1000  to 
2000  c.c. 

As  has  been  mentioned,  the  osmotic  pressure  of  the  gastric  juice 
usually  is  less  than  that  of  the  blood,  and  according  to  H.  Strauss*^ 
it  normally  has  a  freezing-point  depression  of  from  0.36°  to  0.48°. 
Pathologically  it  may  rise  to  0.58,  which  is  the  freezing-point  depres- 
sion of  the  blood.  It  is  of  especial  interest  to  know  that  in  those 
cases  where  osmotic  pressure  approaches  that  of  the  blood,  accord- 
ing to  H.  Strauss,  free  HCl  is  usually  absent.  These  observations 
have  been  confirmed  by  others  (Winter,  S.  Schönborn).  This 
fact  of  itself  would  indicate  that  in  a  stomach  where  the  second 
glandular  function,  the  modification  of  the  ultrafiltrate,  is  arrested, 
the  composition  of  the  gastric  juice  is  more  like  that  of  the  blood 
crystalloids.  We  cannot  omit  to  mention  that  another  fact  is 
opposed  to  this:  in  normal  gastric  juice,  the  NaCl  content  is  nearer 
that  of  the  blood  (0.59  per  cent)  than  in  subacidit3^  Unfortunately, 
I  know  of  no  adequate  data  upon  pure  gastric  juice  from  a  subacid 
stomach,  so  that  for  the  present  the  interpretation  must  remain 
indefinite. 

The  secretion  of  a  juice  with,  free  hydrochloric  acid  from  a  neutral 
fluid,  the  blood,  is  one  of  the  problems  which  offers  especial  diflä- 
culties  to  physiologists.  An  attempt  has  been  made  to  explain  the 
phenomenon  by  the  law  of  mass  action  ^\'ith  the  help  of  carbonic  acid. 
In  my  opinion  colloid  chemistry  offers  analogies  which  permit  an 
unforced  explanation.  We  know  that  neutral  salts  may  he  split  into 
acids  and  bases  by  adsorption  (see  p.  28);  thus,  for  instance,  an  acid 
fluid  with  free  sulphuric  acid  remains  after  shaking  a  solution  of 
potassium  sulphate  ^^ith.  hydrated  manganese  dioxid  (J.  jNI.  Van 
Bemmelen).  With  this  in  mind  we  need  not  be  surprised  at  the 
splitting  off  of  free  HCl  from  a  solution  of  chlorids. 

The  secretion  of  gastric  juice  is  analogous  to  the  secretion  of  acid 
by  plant  roots,  which,  according  to  Baumann  Gu'lly,  hkewise  occurs 


330  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

by  decomposition  of  salts  through  the  adsorption  of  bases  by  the 
pelHcle  of  plant  cells. 

Investigations  of  gastric  secretion,  as  far  as  they  relate  to  physical 
chemistry,  have  hitherto  almost  exclusively  consisted  of  observation 
of  the  osmotic  condition  and  the  electrolyte  concentration;  the  ma- 
terial for  review  is  consequently  as  yet  far  too  limited  for  a  colloid- 
chemical  consideration. 

The  Secretions  Which  Pour  Into  the  Intestines. 

There  are  200  c.c.  of  succus  entericus  daily  secreted  by  the  in- 
testinal glands. 

The  Succus  Entericus  is  usually  hypertonic.  If  we  inject  into 
an  animal  (D'Errico,  chickens,  D'Errico  and  Savarese,  dogs)  a 
hypertonic  common  salt  solution,  the  osmotic  pressure  of  the  succus 
entericus  becomes  still  higher  so  that  its  freezing-point  depression 
may  increase  to  from  0.89°  to  0.99°  (blood  =  0.59°).  This  corre- 
sponds to  our  conception  that  an  ultrafiltrate  may  be  concentrated 
by  the  absorptive  activity  of  the  intestinal  wall.  The  succus  en- 
tericus, like  the  pancreatic  juice  which  pours  into  the  intestine, 
is  almost  neutral.  The  OH  ion  concentration  is  about  10~^  at  18° 
according  to  Auerbach  and  Pick.  Its  alkalinity  is  approximately 
that  of  a  sodium  bicarbonate  solution.  On  account  of  its  higher 
sodium  bicarbonate  content,  pancreatic  juice  has  a  greater  capacity 
to  combine  with  acid  than  has  succus  entericus. 

According  to  H.  Iscovesco  *^  the  colloids  of  the  pancreatic  juice 
are  electropositive;  they  form  with  the  electronegative  colloids  of 
the  succus  entericus  complexes  soluble  in  a  neutral  environment. 

The  Bile:  600  to  900  c.c.  of  bile  are  secreted  per  day. 

The  osmotic  pressure  of  the  bile  is  approximately  that  of  the 
blood;  its  conductivity  is  somewhat  higher.  According  to  H.  Isco- 
vesco, *2  the  colloid  constituents  of  the  bile  probably  have  an  elec- 
tronegative charge. 

The  Kidneys  and  the  Secretion  of  Urine. 

(See  also  p.  409  et  seq.,  on  "Diuretics.") 

We  shall  briefly  review  the  structure  of  the  kidneys  of  vertebrates : 
the  renal  artery  branches  and  in  the  cortex  or  outer  portion  of  the 
kidney  develops  by  the  formation  of  numerous  glomeruli  an  enor- 
mous surface  (Fig.  50).  These  are  knotted  ball-like  branches  of  the 
smaller  arterial  vessels  placed  in  a  vesicle  (Bowman's  capsule).  The 
artery  leaves  Bowman's  capsule,  subdivides  into  capillaries  which 
collect  together  and  form  the  renal  veins.     Bowman's  capsule  has  an 


SECRETION  ÄND  EXCRETION 


331 


outlet  into  the  urinary'  tubules  which  have,  at  first,  a  convo- 
luted course  covered  with  a  thick  layer  of  cells  and  collect  into 
larger  and  larger  tubules  (Fig. 
51),  which  ultimately  discharge 
the  urine  into  the  pelvis  of  the 
kidney. 

The  phenomenon  of  urine  secre- 
tion consists  in  separating  water 
and  crystalloids  from  a  solution 
containing  colloids  and  crystal- 
loids (blood),  though,  usually, 
in  a  much  higher  concentration 
and  with  a  different  proportion 
of  crystalloid  constituents  than 
occurs  in  the  blood. 

Without  going  into  the  vari- 
ous theories  of  urine  secretion, 
we  shall  here  sketch  those  which, 
from  physiological  and  patho- 
logical investigations,  seem  most 
probable.  According  to  this  the 
glomeruli  are  a  filtration  apparatus 
in  which  water  and  crystalloids 
are  filtered  off  while  the  colloids 
are  held  back.     From  this  ultra- 


Con]/olufecf 
Tubule.^ 


Vascular 
Plexus 
(Glomerulus) 

Bowman's 
Capsule 


.-/as  defferens 


P«-" TW/'^  of  Renal  Artery 
Fig.  50.     Glomerular  structure. 


Fig.  51.  Diagram  of  urinary  drainage 
from  Bowman's  capsule.  (From  G. 
Ludwig.) 


filtrate  which  contains  the  crystalloids  in  no  higher  concentration 
than  blood,  later,  possibly  in  the  first  portion  of  the  urinary  tubules, 


332  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

water  and  some  of  the  crystalloids  are  absorbed,  so  that  a  concentrated 
solution,  the  urine,  passes  off. 

In  order  to  give  an  idea  of  the  quantities  of  fluid  which  are  in- 
volved, the  figures  of  H.  Meyer  and  R.  Gottlieb  *  are  reproduced. 
The  blood  contains  about  0.6  per  cent  urea,  the  daily  urine  about 
30  gm.  There  must  thus  be  about  50  hters  filtered  through  the 
glomeruli  and  about  48.5  liters  reabsorbed  by  the  tubules.  Since  in 
24  hours  from  500  to  600  liters  of  blood  flow  through  the  kidneys,  10  per 
cent  of  the  water  of  the  blood  must  be  filtered  off.  This  is  not  at 
all  improbable  when  we  recollect  that  the  afferent  vessels  {vas  afferens) 
have  a  much  larger  lumen  than  the  efferent  (vas  deferens). 

The  filtration  processes  are  relatively  the  least  difficult  to  explain. 
The  criticism  until  recently  offered  that  no  filtration  could  occur 
through  homogeneous  colloid  layers  has  been  disposed  of  by  the  ultra- 
filtration experiments  of  H.  Bechhold.**i  It  must  not  be  insisted 
too  strongly  that  the  phenomenon  in  the  glomeruli  is  a  "filtration" 
since  it  is  evidently  a  process  midway  between  filtration  and  diffusion. 

As  in  the  case  of  every  other  ultrafiltration,  that  in  the  kidney  is 
dependent  upon  the  pressure.  According  to  E.  H.  Starling,  it 
begins  with  an  arterial  pressure  of  at  least  40  mm.  mercury;  below 
this  the  secretion  of  urine  ceases.  In  blood  artificially  diluted  with 
water,  a  minimal  blood  pressure  which  just  maintains  the  circula- 
tion suffices  for  the  secretion  of  urine,^  R.  Gottlieb  and  R.  Magnus 
showed  this  by  permitting  normal  saline  to  flow  continuously  into 
an  animal's  vein.  According  to  Goll  the  urinary  secretion  rises 
and  falls  almost  proportionately  to  the  blood  pressure.  The  experi- 
ment of  D.  R.  Hooker*  furnishes  a  result  in  point.  He  found  in 
the  isolated  dog's  kidney,  that  with  a  constant  perfusion  pressure 
the  quantity  of  urine  formed  was  directly  proportional  to  the  size  of 
the  (artificial)  pulse  pressure.  The  pulsation  of  the  blood  pressure 
plays  an  important  part  in  the  rate  of  filtration.  H.  Bechhold  *'^^ 
compared  the  amount  of  fluid  which  flowed  through  an  ultrafilter 
under  constant  and  under  pulsating  pressure,  and  found  that  in  the 
latter  case  the  filtrate  was  considerably  more  than  in  the  former. 
We  may  imagine  that  with  the  lower  pressure  the  filter  absorbs  the 
fluid  completely  and  it  is  pressed  out  again  with  the  increased  pres- 
sure. This  depends  very  much  on  the  elasticity  of  the  ultrafilter;  if 
it  can  follow  the  variations  in  pressure  very  rapidly,  the  rate  of 
filtration  is  higher  than  for  an  inelastic  filter.     Perhaps  this  will 

1  I  cannot  agree  here  with  Martin  H.  Fischer  (Oedema,  p.  209.)  He  says, 
"Enormous  pressures  are  necessary  to  filter  fluids  through  thin  colloidal  mem- 
branes." This  is  not  correct.  With  suitably  prepared  thin  collodion  membranes, 
a  few  centimeters  of  pressure  suffice  for  ultrafiltration. 


SECRETION  AND  EXCRETION  333 

give  us  the  basis  for  an  understanding  of  certain  changes  in  the 
function  of  the  kidney  in  pathological  and  senile  conditions,  in  which 
the  filtering  membrane  is  inelastic.  From  Avhat  has  been  said  it 
camiot  be  ruled  out,  that  even  in  glomerular  filtration  a  change  in  the; 
salt  content  of  the  filtrate  from  the  blood  may  occur,  since  salts  and 
water  are  not  taken  up  equally  during  swelling  (see  p.  69).  We  can 
thus  more  readily  understand  how  a  salt  solution  hypotonic  towards 
blood  may  flow  from  the  glomeruli,  although  according  to  R.  Burian 
an  isotonic  filtrate  is  always  obtained  upon  ultrafiltration  through  an 
artificial  ultrafilter.  Moreover,  the  difference  in  the  reaction  of 
blood  and  of  glomerular  filtrate  may  contribute  to  the  dilution. 

R.  A.  Gesell  made  a  remarkable  observation.  Repeating  Bech- 
hold's  experiment  he  employed  more  rapid  pulsation  and  lower 
pressure  and  obtained  from  defibrinated  dog's  blood  a  filtrate  richer 
in  globulin  with  the  steady  than  with  the  pulsating  pressure. 

Obviously,  the  glomerular  membrane  is  just  at  the  limit  of  permea- 
bility for  serum  albumin;  albumoses  pass  over  into  the  urine.  We  do 
not  know  whether  the  normal  urinary  colloids  such  as  urochrome  are 
derived  from  the  glomeruli  or  secondarily  from  the  urinary  tubules. 

This  explanation  receives  valuable  support  from  the  investigation 
of  Martin  H.  Fischer.*  As  we  saw  on  page  215  et  seq.,  there  is  in 
the  body  a  dynamic  swelling  equilibrium  for  the  organ  colloids  which 
may  be  regarded  as  constant  for  a  brief  period  in  thirsting  individ- 
uals; in  them  the  secretion  reaches  a  minimum.  With  excessive 
ingestion  of  water,  edema  does  not  by  any  means  occur,  but  the  excess 
of  water  is  excreted  by  the  excretory  glands,  especially  the  kidneys. 
As  we  have  seen  on  page  220,  there  is  a  verj^  narrow  range  of  swelling 
for  the  blood;  as  the  result  of  this  all  water  (free  water  in  contrast 
to  water  of  swelling)  which  is  in  excess  of  what  is  needed  for  the  normal 
condition  of  hydration  of  the  blood  is  filtered  off  bj^  the  kidneys, 
especially  if  the  muscles,  the  main  reservoir,  are  already  saturated 
with  water.  A  most  convincing  proof  that  it  is  not  the  absolute 
quantity  of  water  but  the  swollen  condition  of  the  blood  colloids 
which  is  of  significance  for  the  secretion  of  urine  may  be  found  in 
the  older  experiments  of  E.  Ponfick  and  the  more  recent  ones  of  R. 
Magnus.*^  These  investigators  transfused  a  rabbit  with  the  blood 
of  another  rabbit  so  that  its  blood  was  increased  from  30  to  70  per  cent. 
In  spite  of  this  there  was  no  increase  or  hardly  any  increase  in  the 
excretion  of  urine.     The  same  results  were  obtained  for  dogs  and  rats. 

The  novel  conception  introduced  by  M.  H.  Fischer  *  is  the  fol- 
lowing: blood  rich  in  CO-2  (venous)  has  a  tendency  to  swell,  or  absorb 
water;  blood  -poor  in  CO2  (arterial)  has  a  tendency  to  shrink  or  give 
up  water.     The  blood  passing  to  the  intestines  is  very  rich  in  CO2 


334  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

and,  consequently,  abstracts  water  from  the  intestinal  mucous  mem- 
brane, causing  absorption  from  the  intestinal  lumen.  The  reverse 
process  occurs  in  the  kidneys.  These  are  traversed  by  great  streams 
of  arterial  blood  which  can  give  up  water.  If  this  assumption  is 
correct,  every  increase  in  the  supply  of  oxygen  to  the  kidneys  in- 
creases urine  secretion,  and  every  interference  decreases  it.  Now, 
every  elevation  of  blood  pressure  and  everything  favoring  the 
circulation  in  the  kidneys  signifies  an  improvement  in  the  supply 
of  oxygen  and  vice  versa.  We  might,  however,  wherever  this  occurs, 
just  as  well  attribute  the  change  in  the  secretion  of  urine  to  the 
changed  filtration  pressure  or  rate  of  filtration.  On  this  account  we 
must  select  the  circumstances  which  M.  H.  Fischer  mentions,  which 
always  indicate  an  improvement  or  impairment  in  the  supply  of 
oxygen.  Among  them  is,  "that  blood  poor  in  oxygen  or  rich  in 
carbonic  acid  supplied  to  the  kidneys  for  the  briefest  period  and 
without  even  the  slightest  disturbance  of  the  circulation  otherwise, 
is  inadequate  to  maintain  even  for  the  briefest  period  a  normal 
secretion  of  urine  by  the  kidneys. "  On  the  other  hand,  there  are 
diuretics  which  cause  an  increased  secretion  of  urine  though  accompa- 
nied by  no  change  in  blood  pressure.  M.  H.  Fischer  injected 
hypertonic  solutions  of  various  sodium  salts  (chlorid,  bromid,  iodid, 
sulphate,  tartrate,  phosphate  and  citrate)  and  obtained  with  them  an 
increased  kidney  secretion,  which  stood  in  a  certain  relation  to  the 
lyotropic  action  of  the  salts  involved.  M.  H.  Fischer  explains  his 
results  by  the  fact  that  the  salts  involved  act  by  shrinking  or  de- 
hydrating the  body  colloids,  and  thus  increasing  the  quantity  of  the 
free  or  filterable  water  (see  also  E.  Frey*).  Sugars,  especiall}'' 
dextose,  act  similarly.  M.  H.  Fischer  and  A.  Sykes  thus  explain 
the  thirst  and  diuresis  of  diabetics. 

Some  diuretics,  for  instance,  urea,  cause  no  increased  circulation 
of  blood,  yet  they  still  increase  the  kidney  secretion  as  J.  Barcroft 
and  Brodie  showed.  This  has  been  urged  as  an  argument  against 
the  theory  of  filtration.  In  my  opinion  the  explanation  is  as  follows : 
urea  obviously  causes  deflocculation  of  the  colloids  included  in  the 
kidney  filter.  We  know  from  pages  55  and  162  that  urea  lowers 
the  melting  point  of  gelatin,  decreases  its  rate  of  solidification  and 
facilitates  diffusion  through  gelatin.  All  these  factors  produce  a 
readier  filterability  of  the  water  of  the  blood  and  a  greater  per- 
meability of  the  renal  filter;  to  what  extent  the  reabsorption  of 
water  is  influenced  cannot  be  decided. 

The  experiments  of  E.  Lamy  and  A.  Mayer  *  seem  an  additional 
factor  in  favor  of  the  filtration  theory.  They  tested  the  relationship 
between  the  viscosity  of  the  blood  and  diuresis.  The  following 
tables  definitely  show  that  with  the  diminution  of  friction,    the 


SECRETION  AND  EXCRETION 


335 


secretion  of  urine  increases  and  conversely.  Though  the  relationship 
is  not  as  evident  in  the  second  experiment  as  in  the  first,  it  is  still  quite 
definite. 


Amount  of  urine  secreted 
in  ten  minutes. 

Coefficient  of  friction. 

c.c. 

4.5 

12.5 

Injection  of  20  gm.  cane  sugar  in  40  c.c.  water: 


Amount  of  urine  secreted 
in  ten  minutes. 

Coefficient  of  friction. 

c.c. 

16 
10 
2.5 

1 

8.96 
12.14 
14.89 
14.71 

Injection  of  50  gm.  cane  sugar  in  100  c.c.  water: 


Amount  of  urine  secreted 
in  ten  minutes. 

Coefficient  of  friction. 

c.c. 

8 

3.5 
1.5 
0.5 

11.39 

10.17 
11,75 
13.92 

[The  choice  of  sucrose  for  these  experiments  is  unfortunate  since,  as 
M.  H.  Fischer  and  Woodyatt  have  shown,  sucrose  dehydrates  and 
offers  free  water  to  the  kidney.  Gelatin  and  gum  arable  raise  the 
coefficient  of  friction  without  increasing  urine  as  Starling  has  shown. 
There  is  consequently  no  foundation  for  the  conclusions  of  Lamy  and 
Mayer.     Tr.] 

The  Concentration  of  the  Glomerular  Filtrate. 

The  filtrate  through  the  glomerular  filter  probably  shows  a  concen- 
tration of  crystalloid  constituents  not  much  different  from  that  of  the 
blood  plasma,  although  a  certain  selection  and  redistribution  in  the  rel- 
ative proportion  of  the  crystalloids  by  the  renal  filter  is  conceivable. 
The  blood  shows  a  depression  of  the  freezing  point  of  about  0.56°, 
whereas  the  urine  of  man  in  health  may  vary  between  0.07°  to  3.5°. 

For  a  urine  of  higher  osmotic  pressure  than  the  blood,  the  filtration 
theory  must  look  for  support  to  the  assumption  that  water  must  be  sub- 
sequently withdrawn  from  the  diluted  filtrate  in  the  first  portion  of  the 
urinary  tubules.  The  investigations  of  J.  Demoor  seem  to  me  to  be  of 
especial  importance  for  the  assumption  of  a  subsequent  change  in  con- 
centration in  the  urinary  tubules.  This  author  perfused  hypotonic 
and  hypertonic  salt  solutions  through  the  renal  artery  and  while  he 


336  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

measured  the  variations  of  renal  volume  in  a  Plethysmograph,  he  an- 
alyzed the  fluid  coming  from  the  renal  passages  for  the  same  time. 

The  kidney  perfused  with  a  hypotonic  solution  is  firm  and  pale, 
no  fluid  is  expressed  with  pressure;  the  cells  are  so  swollen  that  the 
lumina  of  the  urinary  tubules  are  almost  occluded  —  typical  swell- 
ing. The  kidney  perfused  with  hypertonic  solution  is  large  and  soft, 
much  fluid  is  expressed  by  pressure,  the  cells  are  shrunken  and  the 
tubules  patent  —  typical  shrinking.     (See  also  R.  Siebeck.) 

In  the  light  of  Martin  H.  Fischer's  theory,  the  explanation  of 
the  concentration  of  the  urine  and  the  tubules  becomes  sim])le. 
We  need  only  recall  that  the  tubules  are  interwoven  with  capillaries 
containing  venous  blood  which  reabsorb  water.  This  conception  is 
supported  by  the  experiments  of  R.  Gottlieb  and  R.  Magnus*  as 
well  as  of  E.  H.  Starling,*  who  found  a  diminished  swelling  of 
the  organ  when  the  supply  of  blood  to  the  kidneys  was  diminished. 

The  greater  concentration  of  the  urine  is  not  the  only  fact  requiring 
an  explanation,  but  also  the  fact  that  the  relative  proportion  of  the 
individual  crystalloid  ingredients  is  different  in  the  urine  than  in  the 
blood  plasma.  [A.  R.  Cushny  in  his  monograph  on  ''  The  Secretion 
of  Urine,"  1917,  divides  the  constituents  of  the  plasma  into  threshold 
bodies  and  nonthreshold  bodies.  Dextrose,  chlorids  and  sodium  are 
excreted  only  when  their  concentration  in  the  blood  exceeds  a  certain 
definite  or  threshold  percentage.  Urea  is  an  instance  of  a  substance 
with  no  threshold.  According  to  Ambard  (Presse  Medicale,  April  25, 
1918),  threshold  substances  are  necessary  for  cell  life.  He  quotes 
Chabanier's  research  showing  that  nonthreshold  substances  have 
a  common  secretory  constant  for  each  individual  and  that  they  all 
seem  to  be  solvents  for  fats.     Tr.] 

Though  in  the  blood,  75  per  cent  of  all  crystalloids  are  inorganic 
molecules  according  to  J.  Bugarszky  and  K.  Tangl,  in  the  urine  they 
comprise  only  from  47  to  66  per  cent  according  to  J.  Bugarszky,  Roth 
and  Steyrer.  In  the  blood  plasma,  0.58  per  cent  are  NaCl,  0.05 
per  cent  urea;  in  the  urine,  on  the  contrary,  the  average  amount  of 
NaCl  is  1  per  cent  in  the  presence  of  more  than  2  per  cent  of  urea; 
in  the  blood  there  is  from  0.08  to  0.12  per  cent  grape  sugar,  but  only 
traces  occur  in  the  urine.  As  has  been  said,  it  is  entirely  possible 
that  even  during  filtration  certain  changes  occur,  so  that  the  relative 
proportion  of  crystalloids  even  in  the  glomerular  filtrate  differs  from 
that  in  the  blood  plasma  though  the  most  important  change  results 
from  the  reabsorption  of  water  in  the  first  portion  of  the  tubules. 

That  concentration  may  be  brought  about  by  swelling  has  already 
been  shown  in  the  classical  example  of  C.  Ludwig  mentioned  on 
page  66.     So  much  water  may  be  withdrawn  from  a  concentrated 


SECRETION  AND  EXCRETION  337 

common  salt  solution  b5^  a  fish  bladder  that  salt  crystallizes  out. 
From  the  investigations  of  F.  Hofmeister  and  Wo.  Ostwald  it  may- 
be concluded  that  the  swelling  of  gelatin-jellies  is  favored  by  some 
salts  in  accordance  with  their  lyotropic  action,  while  other  salts  dimin- 
ish the  swelling  (see  pp.  69  and  70) ;  in  other  words,  according  to  the 
nature  of  the  dissolved  salts,  more  or  less  water  may  be  removed  from 
a  solution  by  the  swelling  of  jellies.  The  extent  to  which  the  individ- 
ual ions  in  mixtures  of  electrolytes  may  be  more  or  less  concentrated 
in  the  solution  which  remains  after  the  swelling  has  received  as  yet  no 
satisfactory  experimental  study.  But  even  Ludwig  sought  such  an 
interpretation  and  sought  by  its  aid  to  explain  the  proportionately 
greater  excretion  of  phosphates  than  of  chlorids  in  the  urine. 

Lagergreen  found  a  negative  adsorption  with  charcoal  and 
kaolin  in  solutions  of  chlorids  of  Na,  K,  NH4  and  Mg;  in  other 
words,  there  resulted  a  concentration  of  the  solution,  whereas  nitrates 
show  a  positive  adsorption;  in  the  case  of  sulphates  the  adsorption  was 
partly  positive  and  partly  negative.  F.  Hofmeister  in  the  case  of 
gelatin  found  that  the  absorption  of  water  and  of  dissolved  substance 
proceeded  independently  of  each  other,  that  the  absorption  of  water 
from  an  NaCl  solution  increased  until  the  concentration  reached  13 
to  14  per  cent,  and  that  when  the  concentration  was  higher  than  this, 
the  absorption  of  water  fell  again.  J.  M.  Van  Bemmelen  demon- 
strated that  potassium  sulphate  is  split  up  by  manganese  dioxid, 
because  K  is  adsorbed  while  SO4  remains  in  solution  (the  solution 
has  an  acid  reaction).  Subsequently,  similar  cleavages  were  demon- 
strated by  M.  Masius  and  L.  Michaelis.  It  is  thus  evident  that 
there  exists  the  possibility  of  a  varying  adsorption  of  salts  (and 
other  substances)  during  swelling;  so  that  an  acid  fluid  (urine  re- 
acts acid)  may  result  from  a  neutral  filtrate.  If  we  now  attempt  to 
apply  these  general  facts  to  the  special  instances  of  the  concentration 
of  the  glomerular  filtrate,  we  shall  see  that  the  most  important 
scientific  support  is  lacking.  Adsorption  ex^Deriments  wdth  renal 
substance  are  especially  necessary.  The  experiments  of  Torald 
Sollmann  may  be  explained  on  the  basis  of  our  hj^Dothesis.  He 
found  that  the  percentage  of  chlorids  in  the  urine  was  increased  by 
nitrates,  iodids  and  sulphocyanids,  and,  on  the  other  hand,  that  it  is 
decreased  by  acetates,  phosphates  and  sulphates. 

No  facts  contradict  the  colloid-chemical  conception  of  urine  ex- 
cretion, but  we  still  lack  the  special  experimental  data  that  should 
support  it.  If  we  recall  that  none  of  the  other  explanations  of 
diuresis  are  in  any  better  position  but  that  they  must  cling  to  ^^tal- 
istic  assumptions,  we  are  compelled  to  accept  the  filtration  theory  as 
the  most  advantageous  until  a  better  one  shall  replace  it. 


338  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Pathology  of  Urine  Secretion. 

As  our  preceding  statements  show,  two  different  functions  of  the 
kidney  may  suffer:  the  filtration  of  the  glomeruH  and  the  concen- 
trating activity  of  the  tubules. 

Filtration  is  deficient  whenever  the  glomerular  filter  is  damaged; 
it  will  also  be  abnormal  if  there  is  nothing  to  be  filtered,  which 
happens  when  an  insufficient  quantity  of  arterial  blood  containing 
free  water  is  supplied  to  the  glomeruli. 

Martin  H.  Fischer  *^  finds  the  chief  cause  of  nephritis  is  an  ab- 
normal increase  in  the  acidity  of  the  kidney  cells.  It  is  the  cause  of 
albuminuria.  According  to  him,  the  acid  dissolves  kidney  protein 
so  that  the  urine  contains  albumin;  it  dissolves  away  the  formed 
elements  (epithelial  cells),  which  are  then  washed  into  the  urine  as 
casts.  Depending  upon  the  (experimental)  conditions,  epithelial, 
granular  or  hyaline  casts  are  formed.  "The  first  change  to  the 
second,  and  these  to  the  third  variety  if  the  acid  concentration  is 
progressively  increased.  Hyaline  may  be  changed  back  to  granular 
casts  if  a  little  salt  is  added  to  the  acid  solution."  (M.  H.  Fischer.) 
M.  H.  Fischer  supports  this  theory  with  numerous  experiments. 

The  increase  in  acidity  may  result  from  a  deficient  supply  of  oxygen 
to  the  kidneys  which  may  be  due  to  a  number  of  different  causes. 
It  may  be  caused  by  deficient  cardiac  activity  of  any  kind,  hemor- 
rhages or  irritation  of  the  vasomotor  nerves,  as  well  as  compression 
of  the  renal  artery  by  a  tumor,  or  interference  with  the  flow  of  blood 
resulting  from  arteriosclerosis  or  embolism.  Congestion  of  the  renal 
vein  must  of  course  have  a  similar  effect.  [Lack  of  exercise  with 
insufficient  breathing  may  induce  acidosis.     Tr.] 

Disturbances  of  renal  function  may  result  directly  or  indirectly 
as  a  result  of  toxic  influences,  such  as  chemical  poisons  and  toxins 
from  infections.  In  such  case  the  oxidation  processes  of  the  renal 
cells  suffer  so  that  renal  edema  results,  and  this  causes  a  compression 
of  the  blood  vessels,  so  that  the  supply  of  arterial  blood  to  the  kidneys 
is  deficient;  a  vicious  circle  is  thus  established. 

M.  H.  Fischer  also  explains  the  action  of  alcohol  and  anesthetics 
in  a  very  plausible  way.  Small  doses  increase  the  excretion  of 
urine  by  increasing  and  strengthening  the  heart's  action,  and  the 
respiratory  frequency  by  vasodilatation;  these  are  all  factors  which 
favor  the  supply  of  oxygen  to  the  blood,  and  in  that  way  the 
formation  of  free  filterable  water.  Caffein  and  digitalis  act  in  a 
similar  way.  Large  doses  of  alcohol,  ether,  chloroform,  chloral,  mor- 
phine, etc.,  on  the  contrary,  bring  about  a  deficiency  of  oxygen, 
causing  a  binding  of  water  by  the  body  colloids  and  thus  a  diminu- 
tion of  the  secreted  urine  (see  E.  Frey). 


SECRETION  AND  EXCRETION  339 

As  the  result  of  these  6olloid-chemical  views,  Martin  H.  Fischer 
successfully  treated  experimental  anuria  (of  rabbits)  by  introducing 
salts  which  counteracted  the  development  of  edema.  Upon  ligating 
the  renal  arteries  of  rabbits  diminished  secretion  of  urine  occurs,  and 
the  kidney  may  be  so  damaged  that  the  anuria  persists.  M.  H. 
Fischer  injected  solutions  of  sodium  phosphate,  sodium  sulphate, 
sodium  chlorid,  after  which  the  edema  receded  and  the  secretion  of 
urine  recommenced.  He  also  obtained  gratifying  results  clinically 
by  administering  hypertonic  solutions  of  sodium  carbonate  and  so- 
dium chlorid  which  is  a  treatment  quite  opposite  to  the  customary  one. 
His  purpose  is  to  hinder  the  accumulation  of  acid  in  the  kidneys. 
The  prohibition  of  violent  exercise,  the  substitution  of  a  vegetarian 
diet  rich  in  alkalis  for  a  meat  diet  and  the  drinking  of  alkaline  min- 
eral waters  is  the  customary  treatment  for  nephritics  and  is  explained 
by  Fischer  on  the  basis  of  his  acid  theory  of  albuminuria.  We 
have  discussed  on  page  239  the  criticism  Fischer's  theory  received. 
Fischer  has  offered  a  new  working  hypothesis  whose  experimental 
discussion  will  be  very  productive,  no  matter  which  side  ultimately 
wins.  [A.  A.  Epstein  has  recently  successfully  treated  edema  in  cer- 
tain types  of  chronic  parenchymatous  nephritis  by  transfusion  and, 
adopting  the  practice  of  Fernand  Widal  and  of  Hermann  StrausS; 
feeding  large  quantities  of  protein,  120-240  gm.  per  day,  in  an  endeavor 
to  increase  the  blood  proteins  which  he  had  found  were  diminished. 
The  increase  in  the  osmotic  pressure  of  the  blood  due  to  the  added 
protein  restored  the  normal  relations  between  tissues  and  blood. 
The  fluid  which  exudes  in  response  to  osmotic  pressure  of  proteins 
should  be  salt  and  water.  Epstein  found  that  such  was  the  com- 
position of  edema  and  effusion  fluids  in  chronic  parenchymatous 
nephritis.     Am.  Jour.  Med.  Sc,  No.  548,  Nov.  1917,  p.  638  et  seq.] 

A  functional  incapacity  of  the  concentrating  activity  of  the  upper 
uriniferous  tubules  becomes  evident  whenever  the  glomerular  fil- 
trate is  not  materially  altered,  and  the  composition  of  the  urine 
approaches  that  of  an  ultrafiltrate  of  the  blood.  As  a  matter  of  fact, 
this  may  be  observed  in  many  cases.  [The  antidiuretic  action  of  the 
extract  of  pituitary  posterior  lobe  and  pars  intermedia  extract  has 
not  yet  been  satisfactorily  explained.  Motzfeld*  concluded,  as  the 
result  of  experiments  on  rabbits,  that  it  is  due  to  a  stimulation  of 
the  renal  vasomotor  system.     Tr.] 

We  saw  that  the  freezing  point  depression  of  the  blood  was  re- 
markably constant  about  0.56°  but  that  for  the  urine  it  varies  be- 
tween 0.07°  and  3.5°. 

A.  VON  KoRANYi*  showed  that  the  "limits  of  accommodation" 
of  renal  function  were  diminished  in  proportion  to  the  severity  of  the 


340 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


renal  disease,  and  that  the  molecular  concentration  of  the  urine  of 
nephritics  approaches  the  molecular  concentration  of  the  blood.  A 
table  from  the  paper  of  A.  von  Koran yi,  Kövesi  and  Roth-Schultz* 
explains  this: 

Freezing  Point  Depression. 


Healthy 

Chronic  interstitial  nephritis 

Chronic  parenchymatous  nephritis . . . 
Subacute  parenchymatous  nephritis . 


Maximal. 


Deg. 
3.5 

0.63-2 
0.68-1.11 
0.75-1.27 


Minimal. 


Deg. 
0.08 

0.12-0.38 
0.36-0.47 
0.53-0.83 


A.  Galleotti*  obtained  from  dogs  suffering  from  phosphorus  and 
sublimate  nephritis,  a  urine  with  a  freezing  point  depression  practi- 
cally identical  with  that  of  the  blood;  Ph.  Bottazzi  and  Onorato* 
obtained  it  likewise  from  dogs  poisoned  with  sodium  fluorid.  An 
experiment  by  these  authors  with  a  dog  suffering  from  cantharides 
nephritis  is,  however,  especially  instructive ;  in  this  case  the  urinif er- 
ous  tubules  are  practically  unchanged  and,  accordingly,  their  activity 
in  concentrating  the  urine  is  practically  uninfluenced. 

If  a  healthy  person  drinks  water  freely,  there  results  a  markedly 
increased  flow  of  urine,  which  in  the  case  of  nephritics  does  not 
occur.  KÖVESI  and  G.  Illyes  examined  the  urine  obtained  through 
ureteral  catheters  from  persons  who  had  a  healthy  and  a  diseased 
kidney.  When  a  great  deal  of  water  had  been  drunk,  there  was  se- 
creted from  the  healthy  kidney  a  large  quantity  of  very  dilute  urine, 
while  from  the  diseased  kidney,  urine  of  average  concentration  was  ob- 
tained. Since  both  kidneys  received  the  same  blood,  the  blood  could 
not  have  been  responsible  for  the  more  dilute  filtrate,  but  the  subse- 
quent dilution  must  have  been  omitted  by  the  kidney  with  the  im- 
paired function.  In  a  diseased  kidney,  not  only  does  the  concentration 
of  the  urine  approach  that  of  the  blood,  but  even  the  amount  of  the 
individual  crystalloids  contained  becomes  similar  to  that  of  an 
ultrafil träte  from  the  blood.  There  are  very  few  investigations  on 
this  subject  and  they  are  chiefly  limited  to  the  chlorids.  Some  data 
supplied  by  Albarran  warrant  our  conclusion.  We  saw  on  page  335 
that  there  is  approximately  twelve  times  as  much  NaCl  as  urea  in 
the  blood,  whereas  in  the  urine  there  is  about  one-half  as  much  NaCl 
as  urea.  Whenever  the  kidney  parenchyma  is  much  diseased,  the 
urea  content  of  the  urine  falls,  and  there  is  a  rise  in  the  amount  of 
chlorids  in  proportion  to  the  amount  of  urea.  However,  the  abso- 
lute quantity  of  NaCl  in  the  urine  diminishes.  In  health  there  is 
approximately  double  the  amount  of  NaCl  in  the  urine  as  in  the 


SECRETION  AND  EXCRETION  341 

plasma;  the  NaCi  content  of  the  urine  of  nephritics  approaches  that 
of  the  plasma.  The  same  fact  seems  to  obtain  for  the  other  crystal- 
loids, including  water. 

Inasmuch  as  the  functionally  inadequate  kidney  continues  to 
ultrafilter  but  ceases  to  regulate  the  ultrafiltrate,  it  likewise  loses 
its  function  as  a  regulator  of  the  entire  organism;  it  is  no  longer  able 
to  maintain  the  "miheu  Interieur."  The  diminished  excretion  of 
crystalloids  by  insufficient  kidneys  causes  an  increase  in  the  crystal- 
loidal  content  of  the  blood  as  was  first  shown  by  A.  von  Koranyi  by 
measuring  the  freezing  point  depression  (cryoscopy). 

The  Result  of  Deficient  Kidney  Function  Upon  the  Organism. 

If  in  animal  experiments  both  kidneys  are  removed,  an  hydremia 
develops  even  though  no  water  is  given,  or  if  the  water  removed 
by  respiration  and  through  the  skin  is  just  replaced.  If  a  ne- 
phritic is  given^  water  ad  libitum,  the  hydremia  does  not  increase 
at  all,  or  when  it  has  reached  a  given  grade  only  to  an  insignificant 
extent;  the  tissues  which  have  been  deprived  of  water  take  it  up 
again. 

There  thus  develops  between  blood  and  tissues  an  equilibrium  in 
which  more  water  enters  the  blood  than  normally.  This  is  not  sur- 
prising since  there  is  in  functionating  kidneys  a  dynamic  equilibrium 
inasmuch  as  there  is  a  current  of  water  from  the  tissues  into  the 
blood  and  from  the  blood  into  the  bladder.  When  the  kidneys  and 
water  maintenance  are  both  deficient,  a  static  equilibrium  occurs, 
which  (to  the  extent  that  we  may  speak  of  it  in  a  living  system) 
represents  the  true  water  equilibrium  between  blood  and  tissues. 
The  question  now  presents  itself:  is  this  equilibrium  conditioned  by 
osmotic  relations  or  by  the  condition  of  swelling  of  the  colloids  in  the 
blood  and  the  tissues? 

The  investigations  of  A.  von  Koranyi,  P.  F.  Richter  and  W. 
Roth,  H.  Strauss,  Kövesi  and  Suranyi  show  that  the  CI  content  of 
the  blood  serum  of  nephrectomized  animals  and  nephritic  men  is 
not  materially  increased;  nor  has  the  electrolyte  content  increased, 
but  on  the  contrary  the  freezing  point  depression  of  nephrecto- 
mized rabbits  rises  from  0.56°  to  0.60°  (normal)  to  from  0.65°  to  0.75° 
(A.  VON  Koranyi).  In  spite  of  the  increased  concentration  of  the 
nonelectrolyte  crystalloids,  the  content  of  water  has  been  increased. 

Let  us  imagine  what  would  happen  if  matters  were  under  the 
sole  influence  of  osmotic  conditions.  Ever  since  Pflüger's  ob- 
servations, we  know  that  metabolic  processes  occur  in  cells.  Con- 
sequently, an  increased  osmotic  pressure  would  have  to  be  present 


342  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

in  the  tissues,  and  this  would  at  first  result  in  an  impoverishment  of 
the  blood  in  water  and  a  proportional  increase  in  molecular  con- 
centration. Gradually  to  the  extent  that  the  metabolic  products 
enter  the  blood,  there  would  occur  a  water  equihbrium  so  that 
finally  the  molecular  concentration  of  the  blood  would  be  increased, 
but  there  would  be  no  change  in  the  content  of  water.  If,  however, 
we  do  find  an  increase  of  the  water,  it  must  follow  from  this  that  the 
osmotic  relations  offer  no  sufficient  explanation  of  these  phenomena, 
and  in  order  to  understand  them  we  are  compelled  to  invoke  the 
condition  of  swelling  in  the  cell  and  blood  colloids. 

The  Urine. 
A.  Normal   Urine. 

Normally  the  urine  contains  no  serum  albumin;  this  does  not 
by  any  means  mean  that  it  is  free  from  colloids.  Even  the  fact  that 
urine  gives  a  moderately  permanent  foam  when  it  is  shaken,  shows 
that  it  contains  colloids.  According  to  H.  Iscovesco*^  these  colloids 
have  an  electronegative  charge. 

Exhaustive  investigations  on  the  total  quantity  of  nondialyzable 
substances  in  the  urine  have  been  undertaken  in  the  laboratory  of 
F.  Hofmeister  (Kumoji  Sasaki,*  M.  Savare,*  W.  Ebbecke*). 
In  the  normal  urine  these  substances  are: 

In  men,  0.87-2.356  gm average  1.44  gm.  per  day. 

In  women,  0.24-0.70  gm average  0.44  gm.  per  day. 

Their  quantity  is  strongly  influenced  by  the  diet,  and  increases 
after  the  ingestion  of  albumin;  it  is  especially  high  on  a  purely  meat 
diet.  Of  less  importance  is  the  work  of  Tamaka,*  who  tried  to 
determine  from  the  viscosity  the  quantity  of  hydrophile  colloids. 
Since  this  was  done  in  undialyzed  urine  which  contains  very  incon- 
stant quantities  of  urinary  salts,  that  influence  the  viscosity  of 
colloidal  solutions  in  some  unknown  way,  the  method  proposed 
cannot  be  utilized. 

L.  Lichtwitz  and  F.  J.  Rosenbach*  showed  that  colloids  could 
be  removed  by  three  equally  useful  methods :  by  dialysis,  by  shaking 
out  in  the  foam  made  with  benzine,  and  by  alcohol  precipitation. 
The  urine  colloids  exert  a  protective  action  upon  colloidal  gold 
(gold  figure  0.69  to  0.81  mg.);  and  in  this  regard  they  are  between 
gum  arable  and  gum  tragacanth  (table  of  R.  Zsigmondy).  Obvi- 
ously they  are  hydrophile  colloids  whose  activity  is  not  diminished 
by  boiling,  evaporation  or  freezing.  By  heating  them  their  protec- 
tive action  is  raised  inasmuch  as  a  finer  distribution  results,  just  as 
occurs  in  the  case  of  gelatin  (L.  Lichtwitz  *2). 


SECRETION  AND  EXCRETION  343 

Other  studies  seek  to  explain  the  nature  of  the  colloid  constituents. 
These  may  be  mucin  (Mörner*-),  chondroitin-sulphuric  acid  and 
nucleic  acid,  which,  according  to  the  studies  from  F.  Hofmeister's 
Institute,  do  not  pass  through  dialyzing  membranes.  Moreover, 
animal  gum  (Landwehr  and  Baisch)  and  a  nitrogen-containing 
complex  carbohydrate  (Salkowski)  probably  belong  to  the  urine 
colloids.  That  the  yellow  coloring  matter  of  the  urine,  urochrome 
of  G.  Klemperer,  is  not  a  colloid  has  been  shown  by  Determeyer 
and  Wagner  *  as  well  as  by  L.  Lichtwitz  and  Rosenbach.* 

The  fact  that  the  surface  tension  of  normal  urine  is  about  10  per 
cent  lower  than  that  of  water  may  also  be  attributed  to  certain  col- 
loid constituents  (see  W.  D.  Donnan  and  F.  G.  Donnan*  as  well  as 
J.  Amann*). 

If  we  approach  the  question  teleologically,  the  purpose  of  the  col- 
loids in  normal  urine  is  obviously  that  the  urine  be  excreted  clear. 
They  prevent  the  formation  of  sediments  within  the  body.  Wolf- 
gang Pauli  as  well  as  H.  Bechhold  and  J.  Ziegler  have  shown 
that  the  presence  of  albumin  decidedly  increases  the  solubility  of 
uric  acid;  and  J.  Lichtwitz  showed  that  the  same  property  was 
possessed  by  the  urine  colloids.^  He  writes  of  a  case  (loc.  cit,  p.  154) 
of  myelogenous  leukemia  in  a  woman: 

"It  (the  urine)  was  clear,  strongly  acid,  free  from  albumin  and  albumoses. 
It  always  had  a  sediment  of  well-developed  uric  acid  crystals,  etc.  The  urine  was 
clear  only  on  October  27;  the  undialyzed  urine  had  no  protective  action  until  the 
urine  of  October  27  was  examined,  then  the  gold  figure  was  about  0.4  c.c." 

Though  the  protective  action  of  the  colloids  has  been  demon- 
strated only  for  uric  acid,  in  my  opinion  it  is  probably  of  signifi- 
cance also  for  other  substances  which  tend  to  sediment. 

A  normal  amount  of  colloid  in  the  urine  is  essential;  if  it  is  de- 
ficient it  may  lead  to  the  formation  of  urinary  calculi,  as  has  been 
shown  by  H.  Schade  (see  p.  272). 

B,  Pathological  Urine. 

The  kidney  behaves  like  a  very  sensitive  membrane.  A  slight 
rise  in  blood  pressure,  even  venous  congestion,  suffices  to  permit  the 
appearance  of  proteins  in  the  urine.  If  the  Iddney  has  been  damaged 
so  as  to  change  the  kidney  parenchyma,  we  are  not  surprised  to  find 
constituents  of  the  blood  mixed  to  a  greater  or  less  extent  with  the 
urine. 

1  As  early  as  1902,  G.  Klemperer  called  attention  to  the  action  of  colloids 
such  as  soaps,  gelatin  and  starch  paste  in  interfering  with  the  precipitation  of 
uric  acid  (G.  Klemperer,  Verh.  d.  Kongresses  f.  inn.  Medizin,  Wiesbaden,  1902). 


344-  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Clinicians  have  paid  more  attention  to  the  serum  albumin  and 
globulin  than  to  the  other  colloidal  constituents.  All  the  usual 
methods  of  detecting  them  depend  upon  the  fact  that  albumin  is 
made  irreversible  by  boiling  or  by  the  addition  of  a  substance  with 
which  it  combines  (sublimate,  picric  acid,  potassium  ferrocyanid, 
etc.),  and  by  a  second  process  (addition  of  nitric  acid  or  other  electro- 
lyte) it  is  flocked  out.  If  the  urine  is  to  be  examined  further,  es- 
pecially for  sugar,  every  trace  of  albumin  must  be  removed  first.  It 
is  frequently  impossible  to  remove  the  final  traces  of  albumin  by 
coagulation  and  flocculation.  In  these  cases  an  adsorptive  sub- 
stance must  be  employed;  the  urine  is  shaken  with  animal  charcoal  or 
diatomaceous  earth  or  a  precipitate  is  formed  in  the  urine  (lead  acetate 
is  added,  filtered  and  the  lead  removed  with  sodium  phosphate). 

The  other  nondialyzable  constituents  also  show  quantitative 
changes,  especially  in  pathological  urine.  They  are  much  increased 
in  lobar  pneumonia  (422  to  488  gm.  per  day)  and  to  an  enormous 
extent  in  eclampsia  (as  high  as  13.84  gm.  per  liter). 

The  determination  of  the  surface  tension  of  the  urine  which  points 
to  certain  constituents  may  in  the  future  become  of  great  impor- 
tance for  diagnostic  purposes.  Though  the  surface  tension  of  normal 
urine  is  about  10  per  cent  less  than  that  of  water  and  is  not  much 
changed  by  either  albumin  or  sugar,  the  salts  of  the  bile  acids  (sodium 
taurocholate  and  sodium  glycocholate)  produce  a  very  definite 
lowering  (as  much  as  40  per  cent  below  that  of  water).  W.  D.  Don- 
nan  and  F.  G.  Donnan  *  found  that  the  degree  of  icterus  ran  parallel 
with  changes  in  the  surface  tension  of  the  urine. 

Fibrin,  nucleoalbumins,  blood  and  blood  pigments,  as  well  as  all 
the  other  organized  constituents  coming  from  the  organism,  are  the 
result  of  local  disease  of  the  kidneys  or  urinary  passages,  and  at  this 
point  they  cannot  be  discussed  at  greater  length.  More  thorough 
studies  of  albumosuria  and  peptonuria  from  a  colloid-chemical  stand- 
point are  much  needed. 

Casts  occupy  an  entirely  distinct  place.  To  a  certain  extent, 
these  are  actually  casts  of  the  uriniferous  tubules;  they  are  spiral 
or  cylindrical  structures  which  occur  in  inflammation  of  the  kidneys 
(nephritis),  whose  form  and  properties  are  of  diagnostic  importance 
(hyaline,  fine  and  coarsely  granular,  etc.).  Casts,  according  to 
M.  H.  Fischer,*^  are  epithelial  cells  of  the  kidney  dissolved  away 
as  the  result  of  the  formation  of  acid.  By  changing  the  concentra- 
tion of  acid,  he  was  able  to  change  hyahne  casts  into  granular  casts 
and  the  reverse. 

Urinary  calculi  have  been  exhaustively  studied  by  H.  Schade* 
(see  p.  272). 


SECRETION  AND  EXCRETION  345 

He  finds  that  they  are  Caused  by  the  clotting  of  fibrin  when  insol- 
uble or  difficultly  soluble  salts  are  simultaneously  excreted.  The 
lamellation  is  caused  by  the  repeated  precipitation  of  fibrin  with  the 
inclusion  of  crystalloid  sediments.  As  many  various  experiments 
showed,  fibrin  has  a  tendency  to  separate  on  surfaces,  so  that  under 
the  conditions  given,  layers  are  formed. 

Sweat  Glands. 

The  daily  excretion  of  sweat  varies  very  widely.  With  the  average 
intake  of  water,  average  atmospheric  temperature  and  at  rest,  it  is 
about  700  c.c.  in  24  hours,  in  a  man  weighing  70  kilos  (according  to 
A.  Schwenckenbecher).  Cramer  found  an  excretion  of  3  liters 
during  a  summer  march,  and  H.  Strauss  was  able  to  establish  a 
loss  of  1/2  to  1  liter  of  sweat  in  a  half  hour  under  the  influence  of 
diaphoretic  procedures. 

The  sweat  glands  are  more  dependent  on  the  influence  of  the 
nerves  than  are  any  other  glands,  yet  it  cannot  be  doubted  that  here, 
too,  ultrafiltration  of  the  blood  plasma  plays  an  important  part, 
since  the  sweat  glands  have  a  knotted  structure  similar  to  that  of  the 
glomeruli  of  the  kidneys.  In  support  of  this  we  have  the  following 
facts:  sweat  contains  only  the  solid  constituents  most  easily  per- 
meable, NaCl  and  urea,  whereas  the  difficultly  diffusible  salts, 
phosphates  and  sulphates  occur  only  in  traces.  The  fact  that 
nitrogenous  products  of  metaboHsm  also  occur,  agrees  with  my 
assumption  that  the  NH2  and  NH3  groups  facilitate  diffusion  (see 
page  411).  The  acid  reaction  is  probably  due  to  the  sebaceous 
glands;  when  the  secretion  is  artificially  increased,  the  sweat  be- 
comes alkahne  (corresponding  to  the  blood  plasma) . 

Milk. 

Of  all  foods,  milk  is  the  most  important;  on  this  account  it  has 
been  investigated  by  food  chemists  and  physicians.  Its  specific 
gravity,  fat  content,  dried  fat-free  residue,  and  even  the  casein  and 
albumin  content  and  the  quantity  of  milk,  sugar  and  salts  (ash) 
have  been  determined,  but  it  is  only  in  the  last  few  years  that  the 
important  part  played  by  the  condition  of  the  colloidal  constituents 
has  been  pointed  out. 

Milk  is  an  aqueous  solution  of  crystalloids  (salts  and  milk  sugar) 
which  contains  the  colloids,  casein  and  albumin,  and  also  an  emulsion 
of  fat. 

Though  the  colloid  constituents  and  the  fat  of  milk  vary  within  wide 
limits  according  to  food,  season,  age,  etc.  (from  5  to  8.585  per  cent),  the 


346  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

water  and  the  crystalloid  content  is  practically  uniform.  G.  Cokn- 
ALBA*  showed  this  by  extensive  investigations  upon  large  dairy 
herds.  The  widest  hmits  for  the  content  of  dissolved  substances 
was  only  from  5.9  to  6.6  per  cent,  whereas  the  variations  were  usually 
from  6.05  to  6.25  per  cent.  From  this  we  must  conclude  that  milk 
is  the  product  of  at  least  two  processes.  One  is  the  result  of  an 
ultrafiltration  of  the  blood  which  yields  an  ultrafiltrate  of  uniform 
water  and  crystalloid  content.  The  colloids  and  fat  are  mixed  with 
this  solution  by  a  second  process. 

The  fat  globules  of  the  milk  (milk  globules)  have  a  diameter  of 
from  0.1  to  22  ijl  averaging  about  3  ß,  yet  it  is  possible  mechanically 
so  to  break  them  up,  that  they  are  no  longer  visible  microscopically; 
according  to  Wiegner  their  average  diameter  is  0.27  fi.  Such  so- 
called  homogenized  milk  is  recommended  as  being  very  easily  digested 
and  it  has  the  advantage  that  the  cream  cannot  be  removed  either 
by  gravity  or  by  centrifuging. 

It  is  known  by  dairymen  that  milk  may  separate  spontaneously 
to  a  certain  extent;  on  standing,  the  cream  rises  to  the  top.  The 
result  is  produced  more  rapidly  and  completely  by  centrifugation; 
though  a  complete  separation  is  not  obtained  even  in  this  way,  we 
have  an  opaque  milk  which  contains  the  finest  globules.  Nor  can  a 
separation  be  obtained  by  filtering  through  the  least  porous  Cham- 
berland  filter.  All  the  fat  globules  can  be  held  back  by  an  ultra- 
filter  which  still  permits  the  complete  passage  of  albumin. 

If  we  wish  to 'make  butter  from  cream,  in  other  words  to  make  the 
milk  globules  unite,  the  cream  must  be  churned,  since  between  the 
individual  globules  aqueous  and  water-soluble  constituents  of  the 
milk  occur  as  partitions,  and  these  partitioning  walls  must  be  broken 
down.  It  is  a  remarkable  fact  that  milk  globules  do  not  dissolve  in 
ether  if  milk  is  shaken  with  it.^  If  milk  fat  were  an  ordinary  emulsion 
such  as  oil  in  water,  the  fat  would  be  completely  removed  by  shaking 
it  with  ether.  From  this  we  conclude  that  the  fat  globules  in  the 
milk  are  surrounded  by  a  pellicle  impermeable  for  ether.  If  potas- 
sium hydrate  or  acetic  acid  are  added,  this  interference  is  removed; 
moreover,  the  fat  may  be  removed  from  the  dried  milk  globules  by 
treatment  with  ether,  and  the  peUicles  will  be  left  behind.  It  is 
absolutely  impossible  to  extract  the  fat  completely  from  homog- 
enized milk  by  shaking  with  ether. 

There  is  quite  an  extensive  but  at  present  practically  useless 
bibliography  (see  Voeltz*)  on  the  pellicles  of  fat  globules.  Especially 
erroneous  were  the  experiments  directed  to  splitting  off  the  pellicles 

1  The  fat  of  human  milk,  with  its  much  larger  quantity  of  albumin,  is  readily 
shaken  out  with  ether. 


SECRETION  AND  EXCRETION  347 

chemically,  since  the  equilibrium  of  the  milk  was  thus  changed. 
The  experimental  arrangements  by  which  Voeltz,  at  least  quahta- 
tively,  estabhshed  the  existence  of  serum  pelhcles,  are  the  most 
fortunate.  No  value  is  to  be  attributed  in  my  opinion  to  the  quan- 
titative results,  since  the  composition  of  the  pelhcles  must  change 
while  they  pass  through  the  layer  of  water.  Voeltz  layered  a 
column  of  milk  about  10  cm.  high  under  a  column  of  water  50  cm. 
high.  The  milk  globules  mounted  through  the  water  and  were  thus 
freed  of  all  water-soluble  ingredients.  The  cream  thus  formed  was 
then  taken  up,  freed  from  fat  and  the  residue  determined.  The 
composition  proved  very  variable  and  qualitatively  contained  the 
ash  and  organized  constituents  of  the  milk  as  far  as  that  could  be 
deduced  from  the  mere  determination  of  ash,  organic  substance, 
Mg,  Ca  and  P. 

By  emulsifying  butter  fat  in  skimmed  milk,  Voeltz  produced 
artificial  pellicles  and  compared  them  with  the  natural  ones. 

In  the  light  of  our  knowledge  of  the  spreading  out  of  colloidally 
dissolved  substances  on  surfaces,  it  ought  not  to  be  very  difficult  to 
explain  the  phenomenon  of  the  pellicles  of  milk  fat  (incorrectly  called 
serum  peUicles).  According  to  G.  Quincke,  a  substance  spreads  out 
at  the  interface  between  two  fluids  (or  a  gas  and  a  fluid)  if  by  this 
means  the  surface  tension  of  the  surface  possessed  in  cormnon  is 
diminished.  Oil  spreads  out,  for  instance,  at  the  interface  between 
water  and  air.  We  know  from  W.  Ramsden*  and  Metcalf*  (see  p. 
34)  that  albumin  and  peptone  separate  out  at  the  interface  between 
water  and  air  as  a  solid.  G.  Quincke  *^  showed  that  a  film  of  gum 
solution  surrounded  each  globule  in  the  pharmacopeal  emulsions. 
H.  Bechhold  *^  bases  his  explanation  of  protective  colloids  which 
prevent  the  flocculation  of  organic  colloids  and  suspensions  on  this 
phenomenon. 

Simple  consideration  shows  that  a  fat  globule  in  an  albumin 
solution  must  surround  itself  with  a  layer  of  albumin.  Since  the 
surface  tension  at  the  boundarj^  surface  of  albumin  or  serum  and  fat 
(0.4  to  1.6)  is  smaller  than  that  of  water  and  fat  (1.6  to  2.4)  the 
albumin  of  milk  must  collect  on  the  surface  of  the  fat  globules. 
An  experiment  of  Ascherson*  shows  the  correctness  of  this  a  priori 
assumption.  Ascherson  emulsified  olive  oil  in  an  alkaline  solution 
of  egg  albumin  and  observed  that  the  oil  droplets  were  surrounded 
by  an  albuminous  membrane.  The  strength  and  composition  of 
the  pellicles  of  the  milk  globules  vary,  of  course,  not  only  with  the 
colloidal,  but  also  with  the  crystalloid  constituents,  especially  the 
salts.  In  passing  through  the  aqueous  layer  (in  Voeltz'  experi- 
ment) the  pellicles  are  again  changed,  and  from  the  investigation 


348  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

above  mentioned  (Ramsden  and  Metcalf),  we  need  not  be  surprised 
that  most  authors  (including  Voeltz)  are  convinced  that  the  pelh- 
cles  surrounding  the  milk  globules  are  solid  membranes.  There  is 
certainly  not  the  slightest  indication  that  this  is  actually  the  case, 
while  the  globules  remain  in  the  milk.  In  addition,  it  must  be 
granted  that  the  composition  of  the  pellicles  vary  with  the  size  of 
the  milk  globules.  Voeltz  believes  that  the  pellicles  of  the  individual 
fat  globules  are  individually  distinct  as  a  result  of  their  origin  (in 
the  mammary  gland).  But  his  own  data  convince  me  that  this 
individual  difference  results  from  purely  physical  causes,  namely, 
the  extraordinary  variations  in  composition  according  as  they 
mounted  quickly  or  slowly  and  according  to  the  size  of  the  fat  globule 
pellicle  examined. 

G.  Wiegnee  developed  an  idea  which  appeared  in  the  first  edition 
of  this  book.  He  compared  the  various  physical  properties  of  ordi- 
nary and  homogenized  milk  and  found  that  with  increasing  sub- 
division of  the  fat  globules  (a  fat  globule  is  subdivided  1200  times 
during  homogenization)  there  is  an  increasing  internal  friction  which 
is  explained  by  the  increased  adsorption  of  casein  by  the  expanded 
surface  of  the  fat  globules.  G.  Wiegner  reckoned,  that,  on  the  basis 
of  Hatschek's  formula  (see  p.  16),  the  thickness  of  the  adsorption 
skins  were  6  to  7  fx/x,  so  that  in  normal  milk  2  per  cent  and  in  homog- 
enized, 25  per  cent  of  the  entire  casein  was  adsorbed  by  the  fat. 

Casein  of  milk  is  itself  insoluble  in  water;  it  is  a  fairly  strong  acid 
which  reddens  litmus  and  displaces  CO2  from  its  salts.  For  in- 
stance, casein  may  be  dissolved  with  the  liberation  of  CO2  if  it  is 
shaken  with  a  suspension  of  calcium  carbonate  in  water.  In  the 
milk  the  casein  is  kept  in  solution  by  lime  salts,  and  it  is  an  old  and 
still  incompletely  solved  problem,  what  sort  of  solution  exists.  If 
milk  is  filtered  through  clay  filters  or  shaken  with  pulverized  clay 
filters  the  casein  is  separated  out  (Hermann  and  Fr.  Dupre  *). 
According  to  some  authors  only  26  to  40  per  cent  of  the  lime  remains 
in  the  whey  after  this  treatment.  If  milk  is  filtered  through  an 
uUrafilter  (Bechhold  **)  without  being  stirred,  the  casein  separates 
out  upon  the  filter,  undissolved.  In  the  case  of  the  powdered  clay 
filter  there  is  obviously  an  adsorption  and  flocculation  of  the  casein 
by  means  of  which  part  of  the  casein-calcium  combination  is  split 
up  just  as  the  salts  of  basic  dyes  are  split  by  textile  fibers  or  wood 
charcoal. 

P.  RoNA  and  L.  Michaelis  *  have  investigated  the  influence  of 
actually  dissolved  and  of  coUoidally  dissolved  lime  by  means  of  the 
"osmotic  compensation  method"  (see  pp.  107  and  108).  They 
dialyzed  whole  milk  against  iron-milk  (milk  from  which  the  proteins 


SECRETION  AND  EXCRETION 


349 


were  removed  by  means  of  colloidal  iron  oxid),  against  whey  (the 
casein  was  removed  by  rennin)  and  against  distilled  water.  Whole 
milk  contained  approximately  about  0.15  per  cent  CaO;  iron- 
milk  about  0.12  per  cent  CaO;  and  whey  only  from  0.04  to  0.06  per 
cent  CaO.  The  amount  of  diffusible  lime  is  accordingly  only  about 
0.06  to  0.07  per  cent  CaO  and  consists  thus  of  about  40  to  50  per 
cent  of  the  entire  lime  contained. 

It  is  remarkable  that  the  iron-milk  contains  more  lime  than  is 
really  diffusible,  so  that  when  casein  is  flocked  out  by  colloidal  iron 
oxid,  calcium  goes  into  true  solution.  But  the  iron-whey  contains 
much  less  phosphoric  acid  than  the  milk  so  that  it  must  have  been 
precipitated  by  the  iron  oxid.  From  this  it  is  evident  that  no 
considerable  quantity  of  calcium  phosphate  is  in  colloidal  solution 
or  the  lime  would  be  retained  with  the  phosphoric  acid  by  colloidal 
iron  oxid.  Evidently  the  lime  exists  to  a  considerable  extent  in 
solution  as  a  slightly  dissociated  casein  salt.  In  favor  of  this  view 
are  the  other  properties  of  milk,  which  are  manifested  when  the  equi- 
hbrium  is  shifted  (depression  of  freezing  point,  conductivity) . 

Albumin.  The  milk  of  various  animals  varies  much  in  the  pro- 
portionate amounts  of  the  chief  constituents;  especially  contrasted 
is  the  relation  between  casein  and  albumin,  as  the  following  data 
show: 


Casein. 

Albumin. 

Cow 

Per  cent. 
3.02 
1.03 

3.20 
4.97 
0.67 

Per  cent. 

0.53 
1.26 
1.09 
1.55 
1.55 

Human 

Goat 

Sheep 

Ass 

The  significance  of  these  differences  upon  the  structure  of  the 
different  organisms  cannot  be  determined  at  present,  though  ac- 
cording to  the  investigations  of  J.  Alexander  and  J.  G.  M.  Bul- 
LOWA,*  the  digestibility  is  influenced  by  this  ratio.  These  investi- 
gators are  of  the  opinion  that  the  reversible  albumin  serves  as  a 
protective  colloid  for  the  irreversible  casein.  They  base  this  on  the 
fact  that  woman's  milk,  which  is  rich  in  albumin,  is  difficult  to 
coagulate  by  acids  or  rennet,  and  that  the  same  condition  obtains 
for  cow's  milk  if  it  is  protected  by  gelatin,  gum  arabic,  albumin  or 
the  like. 

In  the  dark  field,  too,  human  milk  shows  a  finer  division  than 
cows'  milk,  as  was  shown  by  the  investigations  of  J.  Lemanissier,* 
A.  Kreidl  and  Neumann,*  as  weU  as  G.  Wiegner.*    Two  distinct 


350  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

elements  in  cows'  milk  (casein  and  fat)  may  be  definitely  recognized 
in  the  dark  field  though  only  the  fat  globules  can  be  recognized  in 
human  milk.  The  submicrons  are  larger  in  asses'  and  cows'  milk  but 
largest  in  ewes'  milk.  In  boiled  milk  the  submicrons  are  larger  and 
disappear  more  slowly  under  treatment  with  solvents  (potash,  gastric 
juice)  than  in  unboiled  milk,  which  is  an  indication  that  unboiled  milk 
is  more  easily  digestible  than  boiled  milk. 

Human  and  cows'  milk  may  be  distinguished  by  their  ascent  or  rise 
in  strips  of  filter  paper  and  their  diffusion  on  blotting  paper.  For 
instance,  according  to  A.  Kreidl  and  Lenk  *  in  150  minutes  cows' 
milk  ascends  only  2.5  cm.,  while  human  milk  ascends  10.8  cm. 
According  to  Lenk*  this  is  chiefly  due  to  the  viscosity  which  in 
turn  depends  on  the  amount  of  albumin  and  casein  contained.  If 
a  drop  or  two  of  cows'  milk  is  placed  on  blotting  paper,  three  zones 
(fat,  casein  and  solution  of  crystalloids)  are  observed,  whereas  human 
milk  exhibits  but  two  (fat  and  other  ingredients). 

I  refer  to  page  173  et  seq.  for  the  methods  of  examining  milk  and 
dairy  products. 

It  is  necessary  for  completeness  to  mention  the  formation  of  skin 
on  boiled  milk.  The  phenomenon  is  obviously  analogous  to  the 
formation  of  solid  skins  on  dyes  and  peptone  solution  (see  p.  33 
et  seq.). 

Referring  to  the  changed  condition  of  the  surface,  it  might  well 
be  worth  finding  out  whether  the  unavoidable  shaking  during 
prolonged  transportation  damages  milk.  We  learn  from  clinical 
experience  that  raw  milk  is  more  easily  digested  than  boiled  milk. 
[From  the  fact  that  boiled  milk  forms  smaller  curds,  it  offers  a  larger 
surface  and  on  this  account  may  be  more  readily  passed  through  the 
pylorus.  It  may  even  remove  acid  from  the  stomach  by  adsorption. 
Tr.]  This  has  been  attributed  to  the  presence  of  enzymes,  which 
are  destroyed  on  heating,  though  no  one  has  ever  been  able  to  give 
any  proof  of  the  action  of  these  enzymes.  The  most  recent  results 
of  research  indicate  that  considerable  changes  in  condition  are  as- 
sociated with  boiling. 

O.  Grosser  ^  found  by  ultrafiltration  that  the  lime  was  attached 
more  firmly  to  the  milk  colloids  in  boiled  milk  than  in  unboiled 
milk.  The  ultrafiltrates  of  boiled  milk  contained  less  lime  than  those 
of  unboiled  milk.  The  nitrogen  and  phosphorus  content  were 
diminished  in  human  milk,  by  boiling,  but  in  cows'  milk  it  remains 
approximately  the  same.  For  instance,  Grosser  found  the  follow- 
ing quantities  of  CaO  in  ultrafiltrates: 

1  According  to  a  private  communication  (as  yet  unpublished). 


SECRETION   AND  EXCRETION 


351 


Ultrafiltrate  of 
raw  milk. 

Milk  boiled  5  minutes. 

Woman's  milk 

Per  cent  CaO 
0.017 
0.041 

Per  cent  CaO 
0.007 
0.034 

Cow's  milk 

The  length  of  time  the  milk  was  boiled,  influenced  the  results.  The 
ultrafiltrate  of  woman's  milk  after  5  minutes'  boiling  still  contained 
about  one-third,  after  15  minutes  one-tenth,  of  the  original  amount  of 
CaO.     After  boiling  30  minutes  there  were  only  traces  of  CaO. 

That  there  is  hardly  any  difference  between  the  freezing  point 
depression  of  raw  and  of  boiled  milk  is,  therefore,  very  remarkable. 

[McCoLLUM  has  shown  by  his  feeding  experiments  on  rats  that 
dried  milk  contains  both  of  the  essential  food  accessories,  fat  soluble 
A  and  water  soluble  B.     Tr.j 


CHAPTER  XXI. 
THE   NERVES. 

The  nervous  system  began  to  be  investigated  colloid-chemically 
when  the  problem  of  brain  swelHng  and  edema  of  the  brain  was 
formulated  through  Fischer's  theory  of  edema  (see  p.  223).  Various 
attempts  were  made  to  determine  whether  the  swelhng  induced  by 
acids  and  in  the  presence  of  salts  was  analogous  to  the  swelHng  of 
fibrin.  From  the  start  it  was  not  very  probable  that  nervous  tissue 
was  hke  the  latter,  an  individual  protein  substance.  Though  the 
nerve  cell  consists  mainly  of  protein  material,  the  neurolemma  which 
serves  as  insulation  for  nerve  conduction  is  largely  formed  from 
lipoids  which  behave  quite  differently  towards  acids  and  salts. 

Martin  H.  Fischer  and  M.  O.  Hooker  conducted  experiments 
upon  the  swelhng  of  brain  and  cord  in  acids  and  salt  solutions  and 
discovered  that  their  behavior  quite  paralleled  the  behavior  of  fibrin.^ 

For  rehable  data  it  is  important  to  employ  absolutely  fresh  nervous 
tissue  from  normal  animals;  results  of  no  comparative  value  were 
obtained  from  diseased  rabbits  or  animals  which  had  been  chased 
about  before  they  were  killed.  Fischer  justly  criticizes  the  experi- 
ments of  J.  Bauer  and  of  J.  Bauer  and  Ames  who  used  material  6 
to  24  hours  post  mortem;  the  post  mortem  accumulation  of  acids 
rendered  these  experünents  useless  for  the  purpose  of  comparison. 

The  experimental  attempts  of  Barbieri  and  Carbone  to  produce 
swelling  by  injection  of  acids  into  hving  animals  we  must  regard  as 
naive.  The  authors  evidently  overlook  the  fact  that  the  acids  are  dis- 
tributed in  the  organisms  and  that  various  organs  compete  for  the  avail- 
able water;  we  may  expect  swelhng  only  from  local  accumulation  of 
acid.  On  this  account  the  interesting  experiments  of  Klose  and  Vogt 
deserve  elaboration  in  the  direction  of  colloid  research.  The  authors 
found  in  thymectomized  dogs  an  acid  preponderance  locahzed  among 
other  places  in  the  nervous  system  (gray  matter) ;  the  brain  of  a  thy- 
mectomized dog  completely  filled  the  skull,  the  ganghon  cells  were 
swollen.  R.  E.  Liesegang  justly  cautions  anthropologists  against 
attributmg  too  great  significance  to  the  weight  of  brains  (Petten- 

1  This  parallelism  must  not  be  applied  too  generally;  there  are  important 
differences  in  behavior  with  the  salts  of  the  heavy  metals  as  hitherto  unpublished 
experiments  of  H.  Bechhold  show. 

3Ö2 


THE  NERVES  353 

kofer's  brain  weighed  1320  gm.  —  Helmholtz's,  1900  gm.);  for 
sickness,  age,  etc.,  may  produce  considerable  changes  in  a  brain's 
capacity  to  hold  water,  so  that  the  brains  of  men  offer  no  basis  for 
post  mortem  comparison. 

Masuda  suppUed  the  following  figures: 

1st  Man  2nd  Man 

Weight  of  brain 1,291  gm.  1,133  gm. 

Dried  substance 176,36      "  260,30      " 

1  gm.  of  dried  substance  contains  water.  ...       6,14      "  3,55      " 

With  the  same  aqueous  content  as  Brain  II,  Brain  I  would  have 
weighed  767  gm.  With  the  same  aqueous  content  as  Brain  I,  Brain 
II  would  have  weighed  1858  gm.  There  are  increasing  indications 
that  colloid  investigation  is  destined  to  carry  nearer  to  solution  the 
old  problem  of  nerve  irritability. 

Nerve  Irritability  and  Swelling. 

In  considering  muscle  function  we  have  seen  that  it  is  associated 
with  a  certain  condition  of  swelling  (see  p.  294).  The  like  is  also 
true  of  nerves.  The  nerves  lose  their  irritability  when  placed  in  iso- 
tonic solutions  of  cane  sugar  or  other  nonconductors  and  recover  it 
again  when  they  are  placed  in  physiological  salt  solution  (Mathews,* 
E.  Overton  **).  Various  other  neutral  salt  solutions  also  injuriously 
affect  the  irritability  of  nerves  in  accordance  with  a  lyotropic  series. 
The  arrangement  given  by  various  authors  (Mathews,  P.  von 
Grützner)  is  not  as  unequivocal  as  for  muscle.  We  must  remember 
that  the  methods  of  investigation  were  not  the  same  as  for  muscle, 
and  that  the  penetration  of  the  salt  solution  to  the  axis  cylinder 
through  the  lipoid  insulating  layer  was  not  as  uniform.  R.  Höber  *^" 
(p.  307)  summarizes  the  depressant  action  of  ions  on  nerve  irrita- 
bility in  the  f  oho  wing  series: 

Na  <  Li  <  Cs  <  NH4  <  Rb  <  K 
S04<Cl<Br<I 

The  anion  series  is  the  reverse  of  that  for  muscle  irritabiUty. 
We  know  that  in  the  precipitation  of  acid  albumin,  the  lyotropic 
series  is  the  opposite  of  that  for  alkali  albumin ;  we  know  further  that 
contracting  muscle  has  an  acid  reaction;  on  this  account  we  may  with 
some  probability  infer  that  nerve  albumin  is  more  or  less  alkaline 
(negative). 

HÖBER  succeeded  in  rendering  visible  the  changes  which  reveal 
irritability  or  the  absence  of  irritability  of  nerves.  The  sciatics  of 
frogs  still  connected  to  their  gastrocnemii  were  placed  in  isotonic 
salt  solution  until  they  lost  their  characteristic  irritability  to  the 


354  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Faradic  current.  Teased  preparations  of  the  nerves  were  then  suit- 
ably stained.  The  difference  in  the  turgescence  of  the  axis  cylinder 
was  not  only  indicated  by  the  different  intensity  of  the  stain,  but  also 
by  the  fact  that  according  to  the  salts  employed,  the  axis  cylinder 
remained  as  thin  threads  or  were  swollen  to  broad  bands,  and  that  the 
order  in  which  the  swelling  was  affected  agreed  with  the  lyo tropic 
series  characteristic  of  the  loss  of  irritability.  [W.  Burridge,  loc.  cit., 
has  adopted  MacDonald's  view  that  "  in  the  excited  nerve  the  pro- 
tein aggregates  are  agglutinated  to  form  aggregates  of  greater  indi- 
vidual size;  this  change  is  due  to  the  reception  of  a  negative  electric 
charge.  As  a  result  of  this  (K)  salts  are  liberated  which  charge  the 
next  segment  of  nerve  negatively,  and  so  excite  it  and  leave  the  origi- 
nal segment  a  positive  charge.  This  positive  charge  determines  the 
return  of  the  colloid  of  the  original  segment  towards  their  former 
state  of  aggregation."  Burridge  considers  that  it  is  calcium  plus 
the  negative  charge  which  determines  the  coagulative  change,  and 
sodium  plus  the  positive  charge  which  brings  the  colloids  back  to  the 
original  state.  Inhibition,  according  to  this  view,  ''  is  essentially  a 
decalcifying  process."  See  also  page  361.  R.  Lillie,  Science  N.  S., 
Vol.  XIVIII,  No.  1229,  has  offered  an  electromotor  working  model 
of  nerve  conduction  in  the  transmission  of  the  active  state  along  a 
"  passivated  "  iron  wire.     Tr.] 

Cerebrospinal  Fluid. 

Our  most  sensitive  instrument,  the  brain  and  spinal  cord,  is  pro- 
tected by  a  solid  casing,  the  skull  and  spinal  column.  These  latter 
do  not  fit  the  nerve  organ  closely,  but  have  a  certain  amount  of  dead 
space  which  is  filled  with  the  cerebrospinal  fluid.  We  may  add  that 
both  the  bony  case  and  the  central  marrow  are  covered  with  mem- 
branes which  are  connected  by  a  delicate  mesh  work.  This  network 
is  filled  with  fluid  in  which,  to  a  certain  extent,  the  brain  and  cord 
float.  It  is  a  clear,  colorless,  aqueous  fluid  amounting  to  from  60 
to  200  cc.  in  adults.  Besides  a  very  few  formed  elements  it  con- 
tains 1/2  per  cent  of  albumen. 

Several  cc.  may  be  withdrawn,  by  lumbar  puncture,  from  patients, 
without  injury  —  a  procedure  introduced  by  Quincke.  This  pro- 
cedure has  yielded  valuable  physiological  and  pathological  data  of 
scientific  and  diagnostic  importance.  The  fluid  circulates  constantly 
and  slowly.  Under  pathological  conditions  the  fluid  may  be  quickly 
formed  so  that  at  times  several  Hters  may  be  lost  through  a  fistula  in 
a  day. 

There  are  two  opposing  theories  of  cerebrospinal  fluid  formation. 
The  one  assuming  a  secretion  has  most  supporters,  whereas  Mestre- 
ZAT,  who  asserts  an  "elective  filtration,"  has  fewer  followers.     Mes- 


THE  NERVES  355 

trezat's  "  diffusion  governed  by  osmosis  "  expressed  in  our  terminol- 
ogy is  nothing  other  than  ultrafiltration. 

Goldmann's  experimental  data,  subsequently  to  be  mentioned, 
support  this  view,  that  the  choroid  plexus,  the  exceedingly  vascular 
membrane  in  the  ventricles  of  the  brain,  serves  as  an  ultrafilter. 
[J.  McClendon  has  recently  offered  additional  experimental  support 
for  this  view.     Tr.] 

Under  pathological  conditions  the  albumen  content  of  the  cerebro- 
spinal fluid  suffers  very  striking  changes  in  which  the  globuHn  seems 
particularly  affected.  On  account  of  the  minute  amount  of  albumen 
(0.5  per  cent)  the  usual  tests  for  albumen  have  been  refined  though 
none  have  acquired  the  significance  which  has  been  won  by  "the 
colloidal  gold  test"  of  Lange.  It  rests  on  a  modified  chnical  deter- 
mination of  the  "gold  number"  (see  p.  85),  in  other  words  upon 
a  measurement  of  the  protective  action  on  the  gold  hydrosol  by 
cerebrospinal  fluid.  Normal  cerebrospinal  fluid  diluted  with  4  per 
cent  sahne  has  no  effect  in  any  dilution  upon  the  red  color  of  the 
gold  sol.  If  precipitation  occurs  at  any  dilution  it  indicates  a  path- 
ological change.  By  this  reaction,  luetic  affections  of  the  central 
nervous  system  are  detected  when  the  Wassermann  reaction  is  still 
negative  and  there  are  as  yet  no  subjective  changes.  The  reaction 
also  weighs  in  favor  of  diagnosing  luetic  disease  (tabes,  paresis, 
cerebrospinal  syphihs),  when  other  diseases  of  the  brain  and  cord  are 
in  question.  Meningitis,  tumors,  apoplexies  are  distinguished  from 
lues  by  a  shifting  of  the  precipitation  zone.  [Recently  mastic  solu- 
tions have  been  employed  for  the  same  purpose.     Tr.] 

Observations  of  Kisch  and  Runertz  indicate  that  under  certain 
pathological  conditions  (cirrhosis  of  the  liver)  the  surface  tension  of 
the  cerebrospinal  fluid  varies  from  the  normal. 

The  Integument. 

Though  other  portions  of  the  organism  have  greater  or  less  capac- 
ity to  swell,  this  property  is  very  much  limited  in  the  skin,  epidermis, 
hair,  feathers,  scales,  etc.  It  is  also  evident  that  this  Hmitation  is 
essential  for  maintaining  shape  and  for  retaining  water  within  the 
body.  If  the  epidermis  had  unlimited  swelling  capacity  like  gelatin, 
the  fluids  inside  the  organism  would  suffice  to  stretch  the  skin  and  to 
enlarge  it  until  all  shape  was  lost,  and  finafly  the  interior  parts  desic- 
cated. Conversely,  every  natural  atmospheric  dampness  and  every 
rainstorm  still  more  would  cause  an  almost  unlimited  addition  of 
water  from  the  outside;  a  steady  stream  of  water  would  flow  through 
the  skin  and  the  organs,  and  would  be  poured  out  through  the  kidneys. 
If  these  opposing  forces  could  still  establish  an  equihbrium  it  would 
vary  so  much  in  accordance  vnth  meteorological  conditions  that  it  Is 
hard  to  believe  that  the  body  would  have  any  definite  shape. 


356  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

When  we  think  of  aquatic  animals  the  idea  of  an  integument 
which  can  swell  leads  only  to  caricature. 

This  does  not  mean  that  hair,  feathers,  etc.,  cannot  take  up  any 
water.  We  all  know  that  wool  and  feathers  absorb  water  from  the  air 
and  that  in  very  moist  air  they  feel  damp;  in  making  hygrometers, 
human  hair,  which  expands  with  a  certain  degree  of  moisture  and 
contracts  with  dryness,  is  used. 

The  skin  may  be  preserved  thousands  of  years  on  account  of  its 
slight  swelling  capacity.  In  fact,  we  find  the  framework  of  plants, 
cellulose  and  wood,  in  graves  of  animals  or  men  in  addition  to 
bones,  and  usually  hair,  hide  and  leather  articles. 

The  evaporation  of  water  from  the  skin  occurs  not  only  as  the 
result  of  the  secretory  activity  of  the  sweat  glands,  but  there  is  also 
an  '^insensible  perspiration."  As  P.  G.  Unna*^  has  shown  in  con- 
vincing experiments,  this  may  be  either  inhibited  by  fats  or  on  the 
other  hand  increased  by  enlarging  the  surface,  as  with  powders,  coat- 
ing with  gelatin,  collodion,  etc.  (see  p.  416). 

As  in  the  case  of  other  cell  membranes,  the  saturation  of  the  skin 
with  lipoids,  especially  with  lanolin,  is  of  the  greatest  importance. 
W.  FiLEHNE  and  J.  Biberfeld  *  investigated  the  absorption  capacity 
of  clean  keratin  structures  (wool  fibers,  feather  flues  and  human  hair) ' 
as  well  as  such  as  were  saturated  with  lipoids.  As  was  to  be  expected, 
substances  soluble  in  fat  (phenol,  chloroform,  etc.)  were  easily  ab- 
sorbed, whereas  water  and  salts  penetrated  but  slightly.  This  corres- 
ponds with  the  animal  experiments  of  E.  Overton  on  amphibia  and 
of  A.  Schwenkenbecher  *  on  doves  and  mice.  The  almost  complete 
impenetrability  of  the  lipoid-saturated  skin  for  salts  is  of  the  greatest 
importance  to  the  organism  in  maintaining  its  electrolyte  content. 

[Internal  Secretions.  —  W.  Burridge,  by  his  perfusion  experiments 
with  calcium  solutions  of  varying  strength,  has  offered  an  explanation 
of  some  of  the  activities  of  several  hormones.  Pituitary  substance 
apparently  increases  the  response  of  the  uterus  and  the  heart  to  cal- 
cium. Adrenalin  has  a  two-fold  activity  on  the  heart,  a  primary 
depressing  or  surface  action  and  a  secondary  or  deep  augmenting 
action.  During  the  period  of  exalted  cardiac  activity  the  heart  is 
more  responsive  to  calcium  than  previously.  The  factors  involved 
are  the  concentration  of  adrenin,  the  concentration  of  calcium  in  the 
perfusing  solution,  and  the  state  of  the  heart  induced  respectively  by 
adrenin  and  by  calcium.  Excitation  is  viewed  as  a  coagulative 
change  produced  by  calcium  in  certain  colloids. 

The  effect  of  thyroid  secretion  is  similar  to  that  of  alcohol,  causing 
the  circulation  to  be  maintained  on  what  otherwise  would  be  an 
inadequate  calcium  tension  in  the  perfusing  solution.     Tr.] 


PART   IV. 


An  *  after  an  author's  name  refers  to  the  reference  in  the  index  of  names. 

PART   IV. 
CHAPTER  XXII. 

TOXICOLOGY  AND   PHARMACOLOGY. 

In  the  following  chapter  we  shall  chiefly  consider  the  action  of 
foreign  chemical  substances  and  organisms.  If  only  monera  (bac- 
teria, protozoa  and  yeasts)  were  involved,  the  matter  would  be  rela- 
tively simple;  we  might  regard  them  as  suspensions  and  approach 
their  investigation  with  exact  physico-chemical  methods.  It  may  be 
seen  from  the  chapter  on  "Disinfection  and  Agglutination"  that 
these  viewpoints  have  been  successfully  employed. 

Some  of  the  substances  which  are  injurious  to  monera  (bacteria, 
protozoa,  etc.)  we  call  disinfectants,  and  some  preservatives.  One  of 
the  ways  by  which  they  are  tested  is  to  add  the  solution  under  exami- 
nation to  a  suspension  of  bacteria  in  water  or  bouillon  and  observing 
in  what  concentration  growth  is  inhibited.  It  is  obvious  that  this 
action  is  dependent  upon  the  concentration  and  distribution  of  the 
disinfectant  between  bacteria  and  solvent.  Similar  experiments  may 
be  performed  on  higher  aquatic  animals. 

The  problem  becomes  much  more  complex  in  the  case  of  multi- 
cellular organisms,  especially  the  higher  terrestrial  animals,  where 
the  action  may  be  affected  by  the  portal  of  entry.  Water,  so  essential 
to  life,  becomes  a  poison  when  injected  intravenously. 

Depending  on  their  point  of  entrance,  substances  must  pass  through 
membranes,  filters  or  places  with  digestive  ferments  (stomach,  intes- 
tines, etc.).  This  may  either  determine  the  action  and  the  course 
taken  by  the  poison  or  drug,  or  it  may  even  entirely  block  its  en- 
trance. Since  small  intestines  and  colon  send  their  blood  through 
the  portal  vein  to  the  Uver,  substances  which  are  taken  by  mouth  may 
have  no  effect  in  spite  of  being  well  absorbed  if  they  are  strongly 
adsorbed  by  the  liver,  as  happens  in  the  case  of  potassium  salts, 
curare,  etc.  Only  such  substances  are  absorbed  through  the  skin 
as  are  soluble  in  its  fats. 

,  The  action  of  diphtheria  antitoxin  when  injected  intravenously  is 
500  times  stronger  than  when  it  is  injected  subcutaneously  (W. 
Berghaus).     [This  has  been  shown  by  Park  to  be  due  to  its  slow 

absorption  from  the  tissues.     Tr.] 

359 


360  COLLOIDS  IN  BIOLOGY  AND  MEDICINE      , 

In  common  parlance  poisons  are  such  substances  as  are  harmful  to 
warm-blooded  animals  if  they  are  taken  by  mouth  or  are  inspired  even 
in  minute  quantities.  With  few  exceptions  "poisons,"  popularly 
so-called,  are  acids,  alkahs,  various  metalhc  poisons,  CO  and  similar 
substances,  nerve  poisons  which  even  in  minimal  quantities  may 
depress  essential  functions,  e.g.,  strychnine,  atropine,  etc. 

The  actual  poisonous  effect,  the  death  of  the  organism,  is  only  the 
closing  act  of  a  complicated  drama  which  is  enacted  before  our  eyes. 
The  introductory  scenes  are  for  us  no  less  important  since  they 
teach  us  what  phenomena  may  lead  to  a  tragic  termination.  The 
drama  may  end  happily  if  the  concentration  of  the  drug  is  kept 
sufficiently  low  or  suitable  antidotes  are  employed.  From  our  point 
of  view  it  is  impossible  to  separate  Toxicology  and  Pharmacology. 

Next  in  importance  to  the  concentration,  the  most  important 
influence  upon  toxicological  or  pharmacological  action  is  the  dis- 
tribution. The  fundamental  and  essentially  chemical  idea  of  distri- 
bution was  transferred  by  Paul  Ehrlich  to  the  processes  in 
multicellular  organisms.  Minimal  quantities  of  alkaloids  have  such 
intense  action  because,  in  conformity  with  this  view,  they  are  con- 
centrated in  definite  groups  of  nerves.  In  the  treatment  of  infectious 
diseases,  it  is  necessary  to  find  substances  which  will  be  so  distrib- 
uted between  the  hifected  organisms  and  the  infection  producers 
that  the  largest  possible  quantity  becomes  attached  to  the  micro- 
organisms and  the  least  possible  is  attached  to  the  man,  domestic 
animal  or  plant.  It  is  readily  seen  that  the  distribution  may  be 
either  a  chemical  combination,  a  distribution  between  two  solvents 
or  a  variety  of  adsorption.  In  but  few  cases  has  it  been  possible  to 
determine  the  kind  of  distribution  adopted.  H.  Freundlich  has  cal- 
culated from  the  researches  of  W.  Straub  *  that  an  adsorption  equi- 
librium obtains  for  the  distribution  of  veratrine  between  the  heart 
muscle  of  a  marine  snail  {aplysia  limacina)  and  the  bath  containing 
it.     In  the  special  section  we  shall  give  other  examples. 

To  cause  damage,  the  substance  taken  up  must  also  change  the 
affected  organ  of  the  multicellular  organism  and  under  certain  con- 
ditions it  may  be  immaterial  whether  the  change  is  reversible  or 
irreversible.  A  poison  which  renders  the  respiratory  muscles  func- 
tionless  for  only  a  few  minutes  as,  for  instance,  curare,  causes  death 
in  warm-blooded  animals,  though  the  absorption  is  reversible. 
Cold-blooded  animals,  for  instance  the  frog,  may,  on  the  contrary, 
live  for  days  or  even  recover  since  they  are  able  to  breathe  through 
the  skin.  We  must  assume  that  the  organs  most  essential  to  life 
have  special  protection  against  many,  especially  autogenous  toxins. 
This  may  be  a  physical  protection,  in  a  partially  isolating  channel 


TOXICOLOGY  AND  PHARMACOLOGY  361 

about  the  cell  or  cell  group,  or  a  chemical  defense  in  the  sense  of  P. 
Ehrlich,  who  believes  that  in  these  cases  there  is  no  "receptor"  for 
the  poison.     Probably  both  methods  are  involved. 

Although  "distribution"  is  a  very  difficult  and  as  yet  scarcely 
investigated  field,  in  the  study  of  the  organic  changes  due  to  drugs  we 
are  met  largely  by  unsurmounted  difficulties,  since  in  many  cases  no 
external  or  even  histological  change  may  be  recognized.  It  is  our 
hope  that  colloid  research  may  also  prove  of  great  value  here,  since 
it  is  not  only  profound  changes  in  chemical  constitution  which  in- 
terfere with  the  function  of  an  organ,  but  harm  may  Ije  done  even 
by  changes  in  turgescence,  flocculation,  reversible  precipitation  and 
in  fact  even  changes  in  the  size  of  the  particles.  [This  is  the  basis  of 
the  explanation  of  changes  of  muscular  activity  in  response  to  drugs, 
offered  by  W.  Burridge,  to  which  reference  has  been  made.  Tr.] 
More  particularly  than  heretofore,  we  must  observe  the  course  of  an 
injury;  we  shall  have  to  observe  whether  a  permanent  local  change 
occurs  as  in  the  toxic  action  of  most  metals,  or  whether  the  process 
is  reversible  as  in  the  case  of  the  narcotics,  or  whether  after  a  severe 
effect  occurs,  a  moderately  prolonged  after-effect  takes  place  in  the 
organs  involved,  suggesting  an  adsorption. 

Valuable  suggestions  for  such  a  viewpoint  are  offered  by  the 
classical  observations  of  Paul  Ehrlich  on  the  "Oxygen  require- 
ments of  the  body"  and  by  his  study  of  the  histology  of  the  blood 
(P.  G.  Unna*^),  and  W.  Straus's*  researches  on  the  distribution  of 
various  alkaloids  (veratrine,  strychnine,  curarine)  between  heart 
muscle  (of  a  sea  snail)  and  the  surrounding  solution.  [W.  Burridge 
has  shown  that  digitalis  and  strychnine  increase  the  utilization  of 
calcium  by  the  heart.  The  action  of  strychnine  on  nerves  is  explained 
by  a  relative  calcification  of  the  synapse,  facilitating  the  passage  of 
the  nerve  impulse  (see  p.  354).     Tr.] 

Cooperation  of  Indifferent  Substances. 

On  pages  55  and  334,  we  saw  that  the  presence  of  many  substances 
might  either  increase  or  diminish  the  permeability  of  membranes; 
from  this  fact  we  are  justified  in  concluding  that  the  addition  of 
an  essentially  indifferent  substance  may  increase  or  diminish  the 
toxic  or  pharmacologic  action  of  a  substance  by  aiding  or  impeding 
its  arrival  at  the  spot  where  it  is  active.  We  may  in  general  assume 
from  this  that  such  other  substance  is  also  stored  in  the  same  organ. 

We  may  exemplify  this  by  several  observations  (see  0.  Stoffel  *), 
which  must  be  viewed  from  this  standpoint.  Von  Schröder  found 
that  the  diuretic  action  of  caffein  was  increased  by  chloral  hydrate, 
and  yet  that  this  effect  did  not  depend  on  the  ability  of  the  chloral 


362  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

to  paralyze  the  vessels  (i.e.,  increased  blood  supply  to  the  kidneys). 
According  to  Cervello  and  Lo  Monaco,  chloroform  checks  caffeine 
diuresis  when  simultaneously  administered,  but  it  has  no  influence  if  the 
chloroform  effect  precedes  the  caffein.  According  to  Thompson  and 
Walti,  atropine  checks  renal  secretion  and  at  the  same  time  decreases 
the  amount  of  urea.  According  to  H.  Löwe  the  amount  of  urine  se- 
creted was  not  increased  by  the  injection  of  pilocarpine,  the  sugar  re- 
mained unchanged,  the  uric  acid  was  somewhat  increased,  phosphoric 
acid  was  diminished  greatly  and  the  total  nitrogen  to  some  extent. 

It  must  be  emphasized  that  we  are  here  dealing  with  pure  secretory 
activity.  The  action  of  CO2  on  glycosuria  may  possibly  be  attrib- 
uted, according  to  Stoffel,*  to  changes  in  permeability,  whereas 
phloridzin  diuresis  is  much  more  probably  accounted  for  by  a  hin- 
drance to  the  reabsorption  of  the  sugar  formed  in  the  kidneys. 

Since  our  colloid-chemical  knowledge  in  the  realm  of  pharmacologj'' 
and  toxicology  is  extremely  restricted,  we  are  limited  to  the  few 
short  chapters  which  follow.  It  must  be  especially  emphasized  that 
the  most  important  territory,  the  specific  nerve  actions  ^  (see  note 
on  p.  352)  is  colloid-chemically  still  almost  completely  terra  incognita. 

Toxicology  and  pharmacology  study  the  action  of  chemical  and 
physical  influences  upon  organisms,  i.e.,  colloid  structures.  Aside  from 
general  considerations,  the  action  of  suspensions  and  colloids  upon  the 
body  deserves  our  special  attention.  In  the  following  pages,  these 
questions  though  apparently  separated  are  to  a  certain  extent  sys- 
tematically handled,  yet  this  is  upon  superficial  and  not  upon  essen- 
tially scientific  grounds. 

Colloids. 

Pharmacy  and  therapeutics  ever  since  the  classic  age  have  made  con- 
siderable use  of  colloids  and  suspensions,  that  is  of  the  general  colloidal 
properties  of  substances  which  have  absolutely  no  specific  chemical 
action.  I  do  not  refer  to  the  containers  for  medicines,  such  as  gelatin 
capsules  or  wafers,  but  to  the  strongly  adsorptive  properties  of 
colloids  and  suspensions  which  guarantee  a  rapid  action  by  reason 
of  their  enormous  development  of  surface.  Colloids  may  serve  to 
correct  the  action  of  substances  such  as  morphine,  chloral,  aloes,  etc., 

1  We  cannot  always  assume  that  it  is  a  nervous  effect  when,  among  its  other 
actions,  the  substance  involved  acts  upon  the  nerves.  For  instance,  strong 
coffee  aids  digestion.  An  investigation  of  Handovsky,*^  based  upon  observations 
of  A.  Pick,  makes  it  probable  that  the  cause  may  be  found  in  a  specific  property 
of  caffeine,  which  raises  the  internal  friction,  i.e.,  the  ionization  of  albumin.  But 
we  know  from  page  156  that  the  disintegration  of  albumin  starts  with  the  forma- 
tion of  albumin  ions,  and  we  can  consequently  understand  why  caffeine  and  theo- 
bromine, which  is  related  to  it,  favor  the  digestion  of  albumin  by  pepsin. 


TOXICOLOGY  AND  PHARMACOLOGY  363 

or  diminish  their  irritation  of  the  stomach  or  intestines.  Upon  this 
action  rests  the  utihty  of  mucilages  (decoctions  of  salep,  marsh- 
mallow  and  gum  arable),  of  talcum,  etc.,  in  diarrheas  as  well  as  the 
addition  of  gelatin  and  vegetable  mucilages  to  acid  foods  and  fruit 
juices.  In  cases  of  poisoning  with  acids,  alkahs  or  caustic  salts, 
we  are  accustomed  to  employ  as  our  most  important  antidotes,  milk, 
egg  albumen,  gruel  (oatmeal  gruel,  quince  mucilage  or  gum  arable) 
or  emulsions  of  fat  and  of  oil.  Gastric  hypersecretion  also  is  favor- 
ably influenced  by  such  substances  (mucilage  of  gums,  starches, 
bismuth  subnitrate,  talcum,  etc.).  Tappeinee  demonstrated  the 
protective  action  of  such  substances  by  the  following  experiment: 
A  "reflex  frog"  which  is  suspended  with  the  hind  legs  in  an  acid 
solution  withdraws  the  legs  after  a  few  seconds.  If  a  solution  with 
the  identical  amount  of  acid  also  contains  gelatin,  gum  arable,  starch 
paste  or  the  like,  there  will  be  no  reflex  movements.  The  action 
of  adsorbents  in  protecting  against  other  poisons  has  long  been  known. 

In  1830,  the  apothecary  Thouery,  experimenting  on  himself,  took 
without  harm  1  gm.  of  strychnine  (ten  times  the  fatal  dose)  with 
15  gms.  of  charcoal.  The  use  of  charcoal  as  an  antidote  against 
poisoning  though  neglected  in  practice  has  been  mentioned  in  several 
textbooks.  Only  freshly  precipitated  iron  hydroxid  (ferric  hydroxid 
in  water,  antidotum  arsenici)  is  in  general  use  as  an  antidote  for 
arsenic  poisoning,  thanks  to  the  authority  of  Bunsen.  From  the 
earliest  times  greater  usefulness  has  been  accorded  to  the  hydrophile 
colloids  as  gruel  (against  aloes,  cantharides,  Colchicum,  croton  oil), 
milk  or  white  of  eggs  against  mercmy  weed,  glue  solution  against  alum. 

Scientifically  exact  study  of  the  adsorptive  action  of  suspensions 
on  poisons  was  undertaken  only  in  recent  years.  W.  Mechowski, 
Adler,  E.  Zunz  and  L.  Lichtwitz  have  contributed  valuable  re- 
searches on  the  adsorption  of  poisons  (phenol,  strychnine  and  various 
poisons,  arachnolysin)  by  animal  charcoal  which  proved  to  be  in  some 
ways  equivalent  to  kaolin  (bolus  alba),  silicic  acid,  chalk,  diatoma- 
ceous  earth  and  bismuth  subnitrate.  In  practical  toxicology  the 
results  did  not  meet  expectations.  Consequently,  as  a  matter  of 
course,  colloidal  carbon  was  tested.  Sabbatani  actualty  inhibited 
the  toxic  action  of  strychnine  intravenously  by  injecting  simulta- 
neously 6  times  the  quantity  of  colloidal  carbon. 

Adsorption  Therapy. 

The  happy  results  from  adsorption  of  acids  and  poisons  by  char- 
coal, clay,  etc.,  led  to  their  use  even  when  the  acids  and  poisons 
arose  in  the  body  itself.  "Adsorption  therapy."  so-called  by  Licht- 
wiTZ,  was  accordingly  introduced  as  a  therapeutic  procedure.     It 


364  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

can,  indeed,  look  back  upon  an  honorable  and  ancient  past.  Dios- 
coRiDES  recommended  kaolin  (bolus  alba)  as  a  dressing  for  erysipelas, 
poisonings  and  many  other  conditions.  Throughout  antiquity  and 
the  middle  ages  adsorption  therapy  retained  its  reputation  until 
modern  chemistry,  which  could  not  explain  its  action,  delivered  its 
quietus  and  looked  derisively  on  some  nature-therapeutists  who  em- 
ployed it  (Claypastor,  Father  Kneipp).  Stumpf  deserves  credit  for 
having  retested  the  clinical  advantages  from  the  use  of  kaolin,  which 
as  a  therapeutic  measure  had  been  forgotten» 

The  scientific  clinical  apphcation  of  adsorption  is  the  product  of 
the  past  year.  During  the  war  with  its  severe  enteric  infections 
(cholera,  dysentery,  typhoid)  it  has  triumphed  unexpectedly. 

In  addition  to  kaolin  and  sihcic  acid  (prepared  by  the  Gesell- 
schaft für  Electroösmose)  charcoal  has  been  employed  most.  Char- 
coal has  yielded  good  results  in  stomach  conditions  (hyperchorhydria 
and  fermentation) ,  and  also  in  obesity  cures  it  has  been  employed  by 
Lichtwitz  who  removes  by  it  the  important  ingredients  of  the 
chyme  (acids  and  enzymes,  see  p.  329) ,  and  fills  the  stomach  so  as  to 
satisfy  the  distressing  pangs  of  hunger.  Gastric  hypersecretion  is 
also  treated  by  gum  arable,  starches,  bismuth  subnitrate  and  talcum. 

In  the  serious  infectious  intestinal  diseases,  cholera,  dysentery  and 
typhoid,  as  well  as  in  the  gastrointestinal  diseases  of  infants,  kaolin 
and  charcoal  act  not  only  by  adsorption  of  toxins  produced  by  the  in- 
fectious agents  but  also  by  the  adsorption  of  the  bacteria  themselves. 

Finally  we  must  mention  the  original  use  of  kaolin  and  charcoal, 
in  purulent  and  dissection  wounds  as  weU  as  in  catarrhs  (vagina  and 
nose)  and  in  exuberant  carcinomata.  The  action  is  the  same  here  as 
in  the  intestinal  infections.  Naturally,  only  sterile  preparations  are 
employed  in  modern  surgery. 

A  further  advance  has  been  to  impregnate  charcoal  with  drugs  which 
are  then  gradually  yielded.  A  good  effect  was  obtained  in  typhoid  with 
iodine  and  thymol.  A  preparation  of  charcoal  impregnated  with  sul- 
phur (eucarbon)  is  used  as  a  mild  laxative  which  at  the  same  time  re- 
lieves flatulence  by  adsorption  of  bacteria  and  putrefactive  material. 

Dermatologists  employ  powders  extensively  for  a  cooling  effect. 
Obviously  the  powders  absorb  the  water  which  emerges  from  the 
skin  and  as  a  result  of  their  surface  development  accelerate  evap- 
oration; to  a  certain  extent  they  amplify  the  skin  surface.  Good 
results  may  be  obtained  on  burns  and  inflammatory  edemas  with 
thick  layers  of  kaolin.  A  cooling  (febrifuge)  effect  may  be  obtained 
according  to  P  G.  Unna  *^  by  painting  the  entire  body  with  a  thin 
layer  of  gelatin  or  collodion;  this  effect  is  explained  by  the  mag- 
nification of  the  body  surface  (see  p.  355). 


TOXICOLOGY  AND  PHARMACOLOGY  3(J5 

The  intravenous  introduction  of  colloids  has  achieved  great  im- 
portance through  therapeutic  use  of  colloidal  metals  (see  below). 
Other  apparently  quite  indifferent  suspension  colloids  have  a  very- 
powerful  action  when  introduced  into  the  blood  vessels.  One  or  two 
ccm.  of  a  kaolin  suspension  injected  intravenously  into  a  guinea-pig 
induce  a  violent  reaction  with  more  or  less  rapidly  fatal  termination. 
Friedberger  and  Isuneoka  demonstrated  that  this  could  not  be 
attributed  to  emboh  but  that  the  toxic  action  depends  on  the  adsorp- 
tion of  vitally  important  constituents  of  cells  (analogous  to  the 
destruction  of  blood  corpuscles  and,  bacteria  in  vitro,  see  p.  200). 
"Sizing"  the  feet  against  chilblains  and  severe  freezing  is  an  ancient 
household  remedy  which  received  renewed  attention  in  the  winter 
campaign  of  1914-1915.  At  present,  there  is  no  satisfactory  explana- 
tion of  its  action.  [Bayliss  has  recommended  intravenous  adminis- 
tration of  gum  arable  solutions  in  shock  to  increase  the  blood  pressure 
after  hemorrhage.     Delaunay  reports  favorably  on  its  use.     Tr.] 

The  peculiar  effect  of  gelatin  on  the  coagulation  of  blood  is  still 
unexplained.  In  severe  hemorrhages,  purpura  haemorrhagica  and 
hemophilia,  gelatin  is  given  internally  (15  to  20  gm.  daily)  as  well  as 
subcutaneously.  Whether  a  colloid  reaction  occurs,  or  whether  the 
clotting  of  fibrin  is  favored  by  the  calcium  contained  in  the  gelatin  is 
still  an  open  question. 

Colloidal  Metals.i 

If  we  except  the  use  of  finely  emulsified  mercury  in  the  form  of 
blue  ointment,  the  introduction  of  colloidal  silver  by  Crede  *  in  1896 
was  the  first  instance  of  the  employment  of  a  colloidal  metal  because 
of  its  colloidal  nature.  It  was  quite  natural  then  to  test  other  col- 
loidal metals,  mercury,  gold,  platinum,  etc.  The  French  have  been 
especially  industrious  in  the  study  of  the  biological  action  of  colloidal 
metals  (bibhography  given  by  Stodel*),  but  the  comprehensive  in- 
vestigations of  the  Italians,  M.  Ascoli  and  G.  Izar*  as  well  as  E. 
Philippi*  and  Preti,*  anticipated  them  in  showing  that  in  all  prob- 
ability the  action  of  inorganic  hydrosols  in  their  main  features  was 
the  same  as  that  of  the  corresponding  salts  or  of  complex  metal 
salts.  Salts  with  the  cations  concerned  have  in  suitable,  usually 
very  small,  dosage,  an  effect  similar  to  the  action  of  the  hydrosols. 
This  conclusion  was  demonstrated  by  the  experiments  of  P.  Portio  * 
as  well  as  O.  Gros  and  J.  M.  O'Connor,*  but  it  was  first  placed  on  a 

^  A  useful  r6sum6  of  the  methods  of  preparation  and  of  the  properties  of  col- 
loidal metals  may  be  found  in  Th.  Svedberg,  Die  Methoden  z.  Herstellung 
Kolloider  Lösungen  anorganischer  Stoffe  (Th.  Steinkopff,  Dresden,  1910).  The 
older  methods  are  contained  in  the  little  work  of  A.  Lottermoser,  Anorganische 
"kolloide  (Ferd.  Enke,  Stuttgart,  1901). 


366  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

scientific  foundation  by  Th.  Paul.  He  demonstrated  that  colloidal 
preparations  of  silver  split  off  silver  ions  in  aqueous  solution  and  in 
such  quantity  that  the  blood  is  saturated  with  silver  ions  because  it 
can  take  up  very  few  of  them  by  reason  of  the  NaCl  it  contains.  In 
these  investigations  the  interesting  fact  was  disclosed  that  the  various 
colloidal  silver  preparation  behaved  differently  when  diluted  in  aqueous 
solution.  The  Ag  ion  concentration  diminished  when  protargol  is  di- 
luted, it  remains  constant  with  sophol  and  increases  with  lysargin  and 
collar  got.  This  explains  the  difference  in  their  therapeutic  application. 
The  remarkable  fact  that  the  concentration  of  Ag  ions  increases  with 
dilution  is  paralleled  in  complex  substances  as  well  as  in  mixtures  of 
the  weaker  acids  and  their  salts  (increase  of  H  ion  concentration). 
According  to  0.  Goes  the  action  of  silver  nitrate  and  silver  iodid  is 
to  be  attributed  to  the  silver  ions  and  complex  compounds. 

Heretofore  colloidal  metals  and  their  compounds  were  employed 
solely  in  aqueous  solutions;  recently,  however,  metal  organosols  have 
achieved  therapeutic  recognition.  Employing  lanolin  as  a  protective 
colloid,  C.  Amberger  has  prepared  many  colloidal  metal  solutions. 
We  have  interesting  publications  concerning  lanolin  solutions  of 
palladium  hydroxyd  sol  (trademarked  leptynol).  M.  Kauffmann 
employed  it  successfully  in  obesity  cures.  It  acts  as  a  carrier  of 
hydrogen,  increasing  oxidative  processes  which  are  deficient  in  the 
obese.  Certain  psychoses  which  may  be  traced  to  similar  causes 
seem  also  at  times  to  be  favorably  influenced  (W.  Gom). 

Silver  hydrosol  of  all  the  metal  hydrosols  has  been  the  most  carefully 
studied;  the  other  hydrosols  show  great  variations  in  some  respects. 

According  to  G.  Izar,*^  even  the  Macedonians  covered  wounds 
with  silver  plates,  and  in  parts  of  Italy  erysipelas  is  still  treated  in 
the  same  way.  In  the  United  States,  silver  foil  is  employed  in  some 
hospitals  to  seal  open  wounds  (R.  Hunt,  Washington).  Crede  at 
first  employed  Carey  Lea's  colloidal  silver.  Manufacturers  soon 
began  to  make  colloidal  silver  preparations  which  are  sold  under  a 
great  variety  of  names.  Among  the  best  known  are  Argentum  Col- 
loidale  Crede,  which  is  sold  as  Collargol  (von  Heyden).  It  is  pre- 
pared by  the  reduction  of  silver  nitrate  with  ferric  citrate;  a  dextrin 
probably  serves  as  the  protective  colloid.  In  the  case  of  Lysargin 
(Kalle)  a  sodium  lysalbinate  serves  as  protective  colloid.  Electrar- 
gol  and  Argof  erment  are  made  by  electric  pulverization  in  the  presence 
of  a  stabiHzer  (probably  gelatin).  According  to  J.  Voigt  the  linear 
diameter  of  the  particles  in  various  commercial  preparations  varies 
between  14  and  26  fxfx. 

M.  AscoLi  and  G.  Izar  prepare  their  hydrosols  according  to  the 
method  of  G.  Bredig  (pulverization  of  silver,  gold  or  platinum  elec- 


TOXICOLOGY  AND  PHARMACOLOGY 


367 


trodes  by  electric  arcs  under  water).     They  stabilized  some  of  their 
solutions  with  pure  gelatin. 

Since  Crede's  publication  the  Hterature  on  the  action  of  colloidal 
silver  has  become  extremely  extensive,  and  the  results  are  very  con- 
tradictory. It  was  at  first  employed  in  septicemias,  by  some,  with 
professedly  good  results,  and  by  others  without  any  apparent  influ- 
ence. I  have  personally  interviewed  many  practitioners  of  medi- 
cine on  the  action  of  colloidal  silver  and  have  found  among  them 
similar  contradictions.  Some  were  enthusiastic  advocates  of  col- 
loidal silver  therapy  at  first,  but  after  several  failures  dropped  the 
use  of  colloidal  silver  entirely.  E.  Filippi  is  possibly  correct  in  at- 
tributing a  therapeutic  result  only  to  a  single  dose.  [A  similar  ob- 
servation has  been  made  in  connection  with  non-specific  therapy  by 
intravenous  injection  of  typhoid  vaccine  in  rheumatism.  Tr.]  He 
emphasizes  the  decided  difference  in  the  hydrosols  of  different  metals, 
so  that  the  hydrosol  of  the  one  most  suitable  must  be  selected  for  each 
individual  case.  Colloid  silver  is  not  only  said  to  be  active  in  general 
infections,  but  it  has  been  praised  also  in  local  processes,  von 
Oettingen,  who  served  in  the  Russo-Japanese  war,  recommends  it 
heartily  as  a  disinfectant  for  wounds.  [MacDonagh  has  used  col- 
loidal manganese.     Tr.] 


Action  on  Microorganisms. 

The  action  of  colloidal  metals  on  protozoa  (paramecium,  vorticella, 
opalina)  has  been  studied  by  E.  Filippi. 


There  are  killed 


By  Colloidal: 

Silver 

Mercury . . . 
Copper .... 

Nickel 

Palladium . 

Gold 

Platinum.  . 


Paramecium. 


Dilut 
n 

ed  approxi- 
lately: 

450,000 

390,000 

70,000 

24,000 

6,500 

>  4,000 

>  4,000 

Vorticella. 


170,000 

92,000 

36,000 

9,500 

5,200 

>  4,000 

>  400 


It  is  noteworthy  that  the  lethal  threshold  for  salts  of  the  same 
metals  are  very  similar  for  the  same  dilution  and  for  the  same  con- 
tent of  metal. 

Colloidal  silver  has  absolutely  no  effect  on  moulds.  I  found  that 
a  1  per  cent  collargol  solution  which  had  been  loft  unstoppered  was 
covered  after  a  time  with  a  species  of  mould.  Similar  observa- 
tions were  made  by  Filippi  *  with  penicillium  and  aspergillus  in  the 
case  of  different  colloid  metals.  R.  Zsigmondy*^  mentions  that 
moulds  grew  on  his  gold  hydrosol  and  that  the  solutions  were  grad- 


368  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

ually  decolorized  by  them  as  the  gold  precipitated  on  the  mycelia  and 
stained  them  black. 

Earher  investigators  (Crede,  Cohn,  Brunner,  Netter)  ob- 
served only  a  moderate  inhibition  of  growth  (1  :  2000  to  1  :  6000  in 
the  case  of  staphylococcus  aureus)  but  no  destruction  of  the  germs 
by  colloidal  silver.  Recent  studies  of  Cernovodeanu  and  V.  Henri  * 
on  anthrax  bacilli,  B.  coli,  staphylococcus  pyogenes  aureus  and  albus, 
B.  dysenteria,  etc.,  show  a  strong  bactericidal  action  of  silver 
hydrosol  in  test  tubes;  researches  of  Charrin,  V.  Henri  and  Mon- 
NiER-ViNARD  *  show  the  Same  effect  in  the  case  of  B.  pyocyaneus. 
The  size  of  the  particles  in  a  hydrosol  is  of  very  great  importance, 
and  in  fact  the  finely  granular  red  solutions  are  much  more  active 
than  the  coarser  green  ones;  the  former  completely  inhibited  growth 
in  dilutions  of  1  :  50,000  to  1  :  100,000.  [Jerome  Alexander  has 
produced  especially  fine  dispersion  by  a  new  principle.     Tr.] 

Similar  results  were  obtained  for  pneumococci  by  Chirie  and 
Monnier  Vinard.* 

According  to  G.  Stodel,*  colloidal  mercury  in  a  dilution  1  :  132,000 
inhibits  the  development  of  B.  typhi  and  of  staphylococci. 

On  account  of  the  results  obtained  with  colloidal  silver,^  as  well  as 
because  of  the  lack  of  irritating  effect  and  of  toxicity  (it  was  pos- 
sible to  employ  it  in  large  doses  subcutaneously  and  intravenously), 
the  hopes  for  its  therapeutic  action  were  justified.  It  is  remarkable 
that,  instead  of  extensive  especially  planned  animal  experiments, 
clinical  experiments  which  were  at  times  favorable  and  at  times  un- 
favorable have  occupied  the  stage.  The  number  of  times  it  has 
been  employed  clinically  compared  with  animal  experiments  is  com- 
paratively small,  and  it  was  tried  on  many  hopeless  cases. 

The  judgment  of  the  results  depends  largely  on  the  experience  of 
the  chnician  and  is  much  influenced  by  the  subjects;  in  short,  the 
results  hitherto  obtained  lead  to  nothing  definite.  On  this  account 
the  indications  for  use  are  very  inadequate.  It  is  from  the  above- 
mentioned  exhaustive  researches  of  M.  Ascoli  and  G.  Izar  *  that  an 
idea  of  the  mechanism  of  the  action  of  metal  hydrosols  has  been 
obtained.  [Harry  Culver  (Jour.  Lab.  &  Clin.  Med.,  May,  1918) 
found  that  the  gonococcidal  action  of  colloidal  silver  (argyrol,  pro- 
targol,  silvol  and  nargol)  was  diminished  in  vitro  by  aging  the  solu- 
tion by  light  and  by  heat.  He  also  found  that  the  gonococci  became 
resistant  or  adapted  to  -a  particular  preparation  by  growth  in  its 
presence.  This  was  not  a  resistance  to  the  other  colloidal  silver 
preparations  but  specific.  The  importance  of  the  "  protecting  "  sub- 
stance is  evident  from  this  experiment.     Tr.] 

1  According  to  Stodel  also,  colloidal  mercury  is  less  toxic  than  mercury  salts. 


TOXICOLOGY  AND  PHARMACOLOGY  369 

Ferments. 

Ferments  are  much  reduced  in  activity  by  the  salts  of  heavy 
metals.  Since  a  parallelism  has  been  shown  to  exist  between  the 
toxicity  of  colloidal  metals  and  that  of  their  salts,  it  was  expected 
that  the  colloidal  metals  would  exert  a  powerful  action  on  ferments. 
It  is  a  remarkable  fact  that  the  colloidal  metals  proved  to  be  more  or 
less  indifferent. 

The  digestion  of  albumin  by  pepsin,  the  digestion  of  gelatin  by 
trypsin,  the  coagulation  of  milk  by  rennin,  the  cleavage  of  fat  by 
pancreatic  steapsin  and  lipase,  the  fluidification  of  starch  by  pan- 
creatin  and  takadiastase  were  uninfluenced  by  colloidal  silver  (see  M. 
AscoLi  and  G.  Izar*). 

L.  PiNcussoHN  *  examined  the  following  substances  for  their  in- 
fluence on  digestion  with  pepsin:  chemically  prepared  hydrosols  of 
silver,  selenium,  gold,  copper,  bismuth,  mercury  (Hyrgolum)  and 
arsenic;  and  electrically  pulverized  preparations  of  silver,  gold, 
platinum,  mercury  and  bismuth.  In  no  case  was  the  activity  of 
pepsin  increased,  but  it  was  diminished  by  large  doses,  and  least  in 
the  case  of  hydrosols  obtained  by  electrical  pulverization. 

E.  FiLipPi  *  was  unable  to  obtain  any  effect  with  colloidal  metals 
(Au,  Hg,  Cu,  Ni,  Pd)  upon  fermentation  in  the  case  of  yeast,  pepsin, 
trypsin  or  rennin. 

SmaU  quantities  of  silver  hydrosols,  on  the  contrary,  activate  the 
diastatic  ferment  of  the  liver  and  of  the  blood  serum. 

According  to  H.  J.  Hamburger,  the  action  of  staphylolysin,  the 
hemolytic  excretion  of  staphylococci,  is  inhibited  by  collargol.  Ac- 
cording to  W.  Weichardt,  colloidal  platinum  and  palladium  neutral- 
ize fatigue  poisons. 

In  vitro,  C.  Foa  and  A.  Aggazzotti  were  unable  to  demonstrate  any 
action  of  silver  hydrosol  upon  toxins,  but  they  could  if  it  was  in- 
jected into  the  circulation  immediately  after  the  toxin. 

O.  Gros  and  J.  M.  O'Connor  obtained  divergent  results  for  the 
decrease  in  the  strength  of  tetanus  and  diphtheria  toxin  produced  by 
collargol. 

Autolysis. 

In  marked  contradiction  to  the  inactivity  of  silver  hydrosol  on 
most  ferments  is  the  very  considerable  influence  of  metal  hydrosols  on 
the  enzymes  of  autolysis.  If  any  organ,  the  stomach,  liver,  spleen,  etc., 
is  kept,  especially  if  kept  at  body  temperature,  changes  occur  in  it  which 
finally  lead  to  a  softening  and  decomposition  characterized  by  a  more 
or  less  extensive  cleavage  of  the  albumins,  nuclcins,  etc.,  involved. 

This  decomposition  occurs  even  though  the  organ  is  absolutely 


370  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

sterile,  so  that  incidental  bacterial  growths  are  not  the  cause;  it  is 
brought  on  by  a  series  of  different  enzymes  each  of  which  has  a  definite 
function,  and  the  process  is  called  autolysis  or  autodigestion. 

All  the  hydrosols  investigated,  namely,  those  of  silver,  gold,  plat- 
inum, mercury,  palladium,  iridium,  copper,  lead,  ferric  hydroxid 
and  aluminium  hydroxid  have  the  ability  to  assist  autolysis;  M. 
AscoLi  and  his  coworkers,  by  separately  investigating  the  resulting 
products,  were  able  to  determine  the  action  of  the  individual  enzymes. 
For  instance,  the  liver  of  a  recently  killed  animal  was  cut  up  into  small 
pieces  and  passed  through  a  sieve;  it  was  then  diluted  with  water  and 
distributed  in  a  number  of  sterile  vessels  with  1  per  cent  toluol  to  pre- 
vent putrefaction.  In  one  sample  the  albumins  were  immediately 
coagulated,  and  the  total  nitrogen,  as  well  as  the  individual  nitrogen 
fractions,  determined.  Varying  quantities  of  metal  hydrosol  were 
added  to  the  remaining  vessels  and  they  were  kept  for  72  hours  at  37°  C. 

Each  portion  was  then  tested  for 

1.  Total  nitrogen  (according  to  Kjeldahl). 

2.  Nitrogen  (as  monamino  acids). 

3.  Purin-bases  (according  to  Salkowski). 

4.  Albumose-nitrogen  (according  to  Baumann  and  Bömer). 

The  difference  between  the  total  nitrogen  and  the  sum  of  the  other 
values  gave  the  quantity  of  nitrogen  present  as  diamino  acids,  peptone 
and  ammonia. 

In  general,  there  is  an  accelerating  action  on  the  total  autolysis 
as  well  as  on  the  cleavage  of  the  nucleins,  and  the  formation  of 
monamino  acids;  though  there  are  considerable  quantitative  dif- 
ferences between  the  different  hydrosols.  For  instance,  minimal 
quantities  of  Ir,  Hg,  Cu  and  Ag  favor  the  autolytic  process  in 
general,  yet  decidedly  larger  quantities  of  Pb,  Au,  Pt  and  Pd  are 
required  for  this  purpose.  The  same  facts  hold  for  the  formation  of 
monamino  acids.  Small  doses  increase,  while  larger  quantities  of 
hydrosols  interfere  with  the  cleavage  of  nucleins;  however  this 
does  not  hold  true  for  silver,  platinum  and  gold  hydrosols.  Under 
ordinary  circumstances  the  uric  acid  formed  during  autolysis  is 
broken  down  still  further  by  a  uricolytic  ferment;  the  action  of  this 
ferment  is  inhibited  by  silver  hydrosol. 

Though  there  is  no  difference  between  the  action  of  stabilized  and 
unstahilized  silver  upon  autolysis,  such  a  difference  was  noticeable  after 
the  addition  of  defibrinated  blood.  Defibrinated  blood  interferes  with 
the  acceleration  of  autolysis  due  to  unstahilized  silver  hydrosol,  but 
it  does  not  do  so  in  the  case  of  the  stabilized  hydrosol. 

This  observation  is  also  of  great  interest  in  connection  with  the 


TOXICOLOGY  AND  PHARMACOLOGY  371 

theory  of  protective  colloids.  A  priori  we  would  be  justified  in  he- 
lieving  that  no  diflference  exists  between  stabihzed  and  unstabilized 
metal  hydrosol,  but  that  a  stabilization  could  be  produced  by  the 
dissolved  albumins  of  the  hashed  organ  or  of  the  added  blood.  The 
above  example  indicates  the  delicate  adjustments  in  the  mechanism 
of  colloid  protection.  [Different  substances  may  compete  for  the 
protector,  thus  establishing  "  preferential  "  protection.     Tr.] 

It  is  interesting  to  note  in  addition,  that  the  above  investigators 
found  that  minimal  traces  of  prussic  acid,  mercuric  chlorid  and 
Cyanid,  arsenious  acid  and  carbonic  oxid  had  as  toxic  an  effect  on  the 
autolytic  action  of  silver  hydrosol  as  upon  its  abihty  to  split  hy- 
drogen peroxide.  This  process  which  was  exhaustively  studied  by 
G.  Bredig  may  be  made  to  regress  so  that  the  metal  hydrosols  may 
"recover."  The  identical  observation  was  made  by  M.  Ascoli  and 
G.  IzAR  in  respect  to  the  autolysis  by  poisoned  silver  hydrosol. 

Blood:  Hydrosols  of  silver,  lead  and  mercury  have  the  abilit}^  to 
dissolve  red  blood  corpuscles,  whether  the  hydrosols  are  stabilized  by 
gelatin  or  not  (M.  Ascoli  *^).  It  is  also  interesting  to  learn  that  pure 
powdered  silver  causes  hemolysis,  though  this  proceeds  very  slowly. 

The  same  silver  powder  when  repeatedly  used  for  hemolysis  be- 
comes inactive;  serum  inhibits  hemolysis  by  silver.  H.  Bechhold  ^ 
observed  that  a  drop  of  mercury  causes  strong  hemolysis,  which 
serum  did  not  inhibit.  He  also  observed  hemolysis  with  metallic 
lead,  though  this  was  much  weaker  than  in  the  case  of  mercury. 
Metallic  copper  hardens  the  erythrocytes. 

Poisons  do  not  interfere  with  the  action  of  silver  hydrosol. 

It  is  necessary  in  these  effects  to  distinguish  between  the  specific 
activity  of  the  metal  involved  and  the  generic  activity  due  to  the 
development  of  surface.  Hemolysis  is  induced  by  quite  indifferent 
suspensions,  by  kaolin  (Friedberger  and  his  pupils)  as  well  as  by 
barium  sulphate  and  calcium  fluorid  (0.  Gengou).  Such  hemolysis 
is  inhibited  by  serum. 

After  AcHARD  and  E.  Weill,  as  well  as  A.  Robin  and  E.  Weill, 
had  studied  the  influence  of  colloidal  silver,  and  G.  Stodel-  had 
studied  the  influence  of  colloidal  mercury  upon  erythrocyte  pro- 
duction, E.  FiLiPPi,  and  later  Le  Fevre  de  Arric,  carried  these  in- 
vestigations further  and  extended  them  to  other  metal  hydrosols. 
The  results  in  brief  show  that  the  red  blood  corpuscles  are  at  first 
diminished  to  a  greater  extent  than  the  white.  Later  there  is  a  con- 
siderable increase  of  both  red  and  white  blood  corpuscles.     After  the 

1  As  yet  unpublished. 

2  The  fact  that  G.  Stodel  did  not  observe  hemolysis  of  dog's  blood  with  electri- 
cally pulverized  colloidal  mercury  is  remarkable,  and  deserves  further  investigation. 


372 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


prolonged  injection  of  hydrosols  the  red  blood  corpuscles  and  the 
hemoglobin  are  somewhat  increased,  but  there  is  no  noticeable  in- 
crease of  leucocytes.  Silver,  copper,  manganese  and  mercury  prove 
most  active;  platinum,  palladium,  gold  and  nickel  are  much  weaker. 
Identical  results  are  obtained  with  small  doses  of  the  salts  of  these  metals. 

This  does  not  completely  accord  with  the  results  of  0.  Gros  and 
J.  M.  O'Connor,*  who  observed  an  immediate  increase  of  the  polynu- 
clear  leucocytes  just  as  occurs  after  the  introduction  of  any  other 
foreign  substances. 

Very  noteworthy  is  the  observation  of  Filippi,  that  colloidal  silver, 
copper  and  mercury  introduced  into  the  circulation  markedly  in- 
crease phagocytosis. 

The  following  table  obtained  with  slightly  different  experimental 
conditions  on  rabbits  illustrates  this: 

Phagocytosis  of  Aleuron  and  Carmine. 


Normal. 

Ag. 

Cu. 

Hg. 

Pt. 

Per  cent. 
3.12 
5.20 

Per  cent. 
27.50 
37.80 

Per  cent. 

17.80 
40.16 

Per  cent. 

38.00 
16.10 

Per  cent. 
8^20 

Le  Fevre  de  Arric  found,  on  the  contrary,  that  this  assumption 
could  not  be  generalized.  In  experiments  with  silver  hydrosol 
(electrargol)  he  found  in  guinea-pigs  an  increase  in  the  phagocytic 
activity  for  colon  and  typhoid  bacilli;  in  rabbits  there  was  a  diminu- 
tion for  typhoid  bacilli.  In  both  guinea-pigs  and  rabbits  there  was 
an  unfavorable  effect  on  the  phagocytosis  of  pyocyaneus  and  staphylo- 
cocci. 

Metabolism. 

Naturally,  the  processes  occurring  in  the  living  organism  are  far 
more  complicated  than  in  the  individual  organ  elements  or  in  the 
dead  organ.  However,  since  there  were  obtained  from  the  study  of 
autolysis  viewpoints  for  the  action  of  hydrosols  on  the  disintegration 
of  nitrogenous  constituents,  the  investigation  of  the  nitrogen  change 
in  the  living  organism  offered  a  prospect  of  profitable  study  (M. 
AscoLi*  and  G.  Izar,*^  Filippi  and  Rodolico). 

For  this  purpose  bitches  were  fed  entirely  on  bread  made  from 
wheat  or  rye  flour.  The  total  nitrogen  in  the  feces  was  determined, 
and  in  the  urine  the  total  nitrogen,  the  urea  nitrogen  and  the  uric  acid. 
In  a  previous  series  of  experiments  with  men,  like  determinations  were 
made  (excepting  of  the  N  of  the  feces)  as  in  the  experiments  under- 
taken on  rabbits  by  E.  Filippi  and  Rodolico.  Metal  hydrosols  were 
administered  intravenously.     The  results  were  concordant. 


TOXICOLOGY  AND  PHARMACOLOGY  373 

The  result  of  the  experiment  was  as  follows:  unstabilized  silver 
hydrosol  (prepared  according  to  G.  Bredig)  as  well  as  collargol  had 
no  action  in  small  doses.  Silver  hydrosol  (prepared  according  to  G. 
Bredig),  stabilized  with  gelatin,  increased  the  nitrogen  metabolism; 
the  nuclein  metabohsm  was  chiefly  affected  since  there  resulted  a 
decided  increase  in  the  elimination  of  uric  acid  in  the  urine.  Sil- 
ver hydrosol  stabilized  by  gelatin  has  a  more  powerful  action  than 
the  corresponding  quantity  of  silver  nitrate,  silver  thiosulphate  or 
silver  albuminate,  which  exert  a  qualitatively  analogous  action. 
On  the  other  hand,  the  N  elimination  in  the  feces  is  decreased. 
Mercury  and  lead  hydrosols  have  a  similar  effect,  differing  only  in 
the  time  curve.  Large  quantities  of  collargol  also  increase  the  uric 
acid  excretion. 

Temperature  Curve. 

The  injection  of  a  few  cubic  centimeters  of  silver  hydrosol  causes 
a  rise  of  temperature  of  varying  but  usually  brief  duration  (M. 
AscoLi  and  G.  Izar*);  on  the  other  hand,  the  unstabiUzed  hydrosols 
have  no  observable  effect  on  temperature  (Bourgougnon*).  This 
corresponds  with  the  observations  on  autolysis  described  above. 

Distribution. 

Finally,  we  must  inquire,  what  becomes  of  the  injected  silver 
hydrosol.  This  has  already  been  investigated,  at  least  as  far  as  con- 
cerns collargol  injected  intravenously.  G.  Patin  and  L.  Roblin  * 
found  it  chiefly  in  the  liver  but  to  a  less  extent  in  the  kidney.  They 
contend  that  there  occurs  a  concentration  and  gradual  excretion 
through  the  kidneys.  S.  Bondi  and  A.  Neumann  showed  that  col- 
largol as  well  as  other  indifferent  suspensions  (India  ink,  fat)  dis- 
appear from  the  circulation  within  1/2  to  1  hour  after  intravenous 
injection  and  are  temporarily  deposited  ift  the  liver,  bone  marrow 
and  spleen.  It  is  the  star  cells  of  von  Kupffer  which  chiefly  take  up 
these  suspensions. 

J.  Voigt  contributed  especially  accurate  researches.  He  traced 
the  fate  of  the  stored  silver  in  the  more  important  organs  bj^  examin- 
ing microscopic  sections  in  the  ultramicroscope.  Of  his  findings  let 
us  emphasize  particularly  that  it  made  a  difference  in  the  distribution 
of  the  silver  in  the  individual  organs  whether  the  animal  was  over- 
whelmed by  a  single  large  quantity  of  silver  solution  or  smaller 
repeated  doses  were  injected.  There  were  definite  differences  in  the 
pictures  obtained  with  different  colloidal  metals  and  metallic  com- 
pounds.    According  to  personal,  hitherto  unpublished  communica- 


374  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

tions  from  J.  Voigt  the  silver  is  precipitated  at  the  site  of  injection 
after  intramuscular  injections  and  in  the  peritoneum  after  intra- 
peritoneal injections,  whence  it  is  gradually  transported  to  the  internal 
organs.  It  is  still  an  open  question  whether  the  transportation  is 
purely  mechanical  or  results  from  solution  and  reprecipitation. 

Therapeutics. 

It  is  obvious  from  the  preceding  statements  that  metal  hydrosols 
may,  from  very  different  causes,  exert  a  therapeutic  action.  In  in- 
fectious processes  we  may  imagine  that  there  is  a  direct  action  on 
the  excitants  of  infection;  although  this  may  be  due  to  an  indirect 
action  inasmuch  as  the  hydrosol  stimulates  the  formation  of  anti- 
bodies and  phagocytosis,  or  it  may  injure  the  infecting  organisms  by 
intensifying  metabolic  changes  in  some  way. 

In  view  of  G.  Bredig's  experiments  on  the  catalytic  action  of  col- 
loidal metals,  a  catalytic  action  of  metal  hydrosols  which  produces 
effects  similar  to  the  ferments  in  the  living  organisms  has  been 
frequently  suggested.  Personally,  I  prefer  to  leave  undecided 
whether  such  an  expression  as  "catalytic  action"  has  any  real  mean- 
ing in  this  connection  or  whether  it  is  nothing  but  an  empty  word. 

We  shall  merely  mention  here  the  experiments  with  colloidal  mer- 
cury, which  has  been  chiefly  used  in  syphilis  and  shows  a  specific 
action  similar  to  that  of  other  mercury  preparations. 

Animal  Experiments. 

In  the  case  of  silver  hydrosol  there  exist  many  experiments  of 
C.  FoA  and  A.  Aggazzotti.*  They  infected  rabbits  with  staphyl- 
ococci and  after  an  hour  injected  30  cc.  of  a  red  silver  hydrosol, 
repeating  this  several  times.  In  this  way  they  delayed  the  death 
of  the  animal  from  1  to  3  days  but  recovery  was  not  brought  about. 

In  infections  with  diplococci  and  typhoid  (in  dogs)  the  animals 
could  be  kept  alive  with  injections  of  silver  hydrosols.  In  the  latter 
instance  this  was  even  possible  when  the  silver-hydrosol  injection 
was  given  in  doses  of  5  cc.  intraperitoneally  as  late  as  12  to  24 
hours  after  the  injection  of  the  microorganisms. 

The  same  authors  found  that  silver  hydrosol  has  no  effect  on 
toxins  in  vitro,  whereas  it  inhibits  the  toxicity  if  it  is  injected  imme- 
diately after  the  toxin.  From  this  they  concluded  that  silver  hydro- 
sol activates  the  oxidizing  ferments  of  the  body. 

Charrin,  V.  Henri  and  Monnier-Vinnard  *  speak  very  guard- 
edly concerning  their  therapeutic  results,  and  characterize  them  as 
"very  promising."     Chirib  and  Monnier-Vinnard*  experimented 


TOXICOLOGY  AND   PUARMACOLOdY  375 

with  pneumococci  on  white  rats  and  mice.  They  ol>tained  at  times 
a  retardation  of  the  disease  process  and  in  individual  instances  they 
allege  a  cure  by  means  of  silver  injections. 

Clinical  Experiments. 

I  shall  pass  over  the  majority  of  experiments  which,  because  of 
their  limited  scope,  are  without  significance  and  frequently  contra- 
dictory, and  shall  only  regard  such  results  as  are  unimpeachable. 
To  all  appearances,  only  experiments  performed  with  a  stabilized 
silver  hydrosol  have  practical  value. 

The  use  of  silver  hydrosol,  as  collargol,  in  septicemia  and  pyemia 
is  most  frequent  and  best  known.  It  is  usually  used  as  an  intra- 
venous injection,  at  times  as  an  ointment  or  an  enema.  If  the 
numerous  case  histories^  are  reviewed,  two  phenomena  are  prom- 
inent: the  fall  in  temperature  and  the  subjective  improvement 
of  the  patient  which  follow  several  hours  after  the  application  is 
given.  In  contrast  to  this  it  is  hardly  possible  to  determine  to 
what  extent  the  disease  process  is  influenced.  The  effect  of  silver 
hydrosol  on  pneumonia  has  been  studied  most  thoroughly.  G. 
Etienne*  and  J.  Cavadias  obtained  good  results;  the  rapid  defer- 
vescence is  also  the  most  significant  fact  here.  G.  Izar*^  treated 
28  cases  of  pneumonia  with  silver  hydrosol  and  several  with  plati- 
num and  iridium  hydrosol;  no  difference  was  noted  between  the 
Ag,  Pt  and  Ir.  These  thoroughly  studied  cases  gave  the  followdng 
results:  the  course  of  the  pneumonia  process  seems  in  general  to 
have  been  favorably  influenced  though  it  was  hardly  possible  to  at- 
tribute this  to  a  specific  action  upon  the  infectious  process,  but 
rather  to  the  amehoration  of  the  symptoms.  As  in  the  case  of 
healthy  individuals,  in  a  pneumonia  patient  a  rise  of  temperature, 
which  reaches  its  maximum  in  about  4  hours,  follows  the  injection 
and  this  is  followed  by  a  severe  rigor,  which  is  succeeded  by 
profuse  sweating  and  a  rapid  temperature  fall,  "critical  in  character, 
however,  it  cannot  be  termed  a  crisis."  The  subjective  improvement 
of  the  patient  is  characteristic  of  the  action  of  silver  hydrosol. 

The  brief  period  of  oppression  and  anxiety  which  accompanies 
the  rigor  is  succeeded  when  the  temperature  falls  ]\v  a  feeling  of  well- 
being  or  euphoria.  Cardiac  and  renal  functioning  are  not  affected, 
nor  is  there  any  action  on  the  course  of  the  pnemnonia  process  as  far 
as  may  be  determined  from  a  change  in  the  excretion  of  chlorids. 

1  A  very  complete  bibUography  is  given  by  Weissmann,  Über  Kollargol. 
Therapeut.  Monatsh.,  Aug.,  1905. 

Mentioned  by  Iscovesco,  Presse  Mddicale,  May  S,  1907. 


376  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

G.  IzAR  reaches  the  conclusion  that  "the  regular  use  of  the  injections 
shortens  the  course  of  the  infection  and  seems  to  make  it  more  favor- 
able." 

It  was  mentioned  at  the  outset  that  the  number  of  infectious 
diseases  in  which  silver  hydrosols  as  well  as  other  metal  hydrosols 
were  employed  is  very  great,  and  the  opinions  of  the  results  very 
divergent;  silver  hydrosol,  and  at  times  platinum-hydrosol,  have  been 
employed  in  inflammatory  rheumatism  and  erysipelas,  in  typhoid 
and  para-typhoid,  in  appendicitis,  furunculosis,  phlegmons,  anthrax, 
cerebrospinal  meningitis,  and  scarlatina,  dysentery  and  diphtheria, 
etc.  As  in  the  case  of  the  diseases  previously  described,  it  affects 
the  temperature  curve  though  at  times  only  temporarily,  and  there 
is  frequently  no  influence  on  the  patients  subjectively. 

I  have  not  as  yet  discovered  in  the  literature  any  published  cases 
of  the  use  of  silver  hydrosols  in  tuberculosis;  if  they  exist  they  are 
probably  isolated  instances.  The  reader  may  well  get  the  impression 
that  there  do  not  exist  for  most  diseases  such  thorough  studies  as  G. 
Izar's  *^  in  pneumonia,  and  that  on  this  account  th  ^  records  of  metal 
hydrosol  therapy  are  incomplete. 

Mercury. 

Mercury  has  been  used  for  centuries  in  syphilis.  Since  metallic 
mercury  as  such,  as  well  as  in  the  very  finely  emulsified  form  of  blue 
ointment,  is  absorbed  by  the  organism,  there  is  no  reason  for  expecting 
a  very  marked  difference  to  result  from  the  colloidal  solution. 

The  chemical  firm  of  von  Heyden  manufacture  a  mercury 
hydrosol  called  Hyrgolum  and  a  mercurous  chlorid  hydrosol  called 
Calomelol,  which  may  also  be  employed  for  inunctions. 

Sulphur. 

For  some  time  a  water-soluble  sulphur  hydrosol  has  been  intro- 
duced into  medicine  and  employed  in  skin  diseases.  Its  action 
depends  on  the  method  of  introduction  since  sulphur  is  reduced  to 
the  highly  toxic  hydrogen  sulphid  in  the  organism.  The  lethal  dose 
for  a  rabbit  weighing  1  kilo,  according  to  L.  Sabbatani,  is  0.0066  gm. 
of  colloidal  sulphur  intravenously  (death  is  immediate) ,  whereas  death 
occurs  only  after  several  hours  when  0.25  gm.  is  introduced  into  the 
alimentary  tract.  The  action  also  depends  on  the  kind  of  animal;  dogs 
are  much  less  sensitive  to  sulphur  than  other  experimental  animals. 

The  reduction  and  consequently  the  toxicity  depends  on  the  physi- 
cal condition;  it  is  most  intense  in  colloidal,  less  in  amorphous,  and 
least  in  crystalline  sulphur.     Moreover  the  toxicity  is  directly  pre- 


TOXICOLOGY  AND  PHARMACOLOGY  377 

portional  to  the  dispersion.     Joseph  recommends  sulphur  hydrosol  in 
diseases  of  the  skin. 

Phosphorus,  Arsenic,  Antimony. 

Of  all  the  complicated  phenomena  caused  by  these  three  sub- 
stances in  different  doses,  there  is  only  one  which  can  be  considered 
coUoid-chemically.  Phosphorus,  arsenic  and  antimony  greatly  influ- 
ence metabolism.  Whereas  arsenic  and  arsenic  salts  inhibit  liver 
autolysis  even  in  small  doses,  minimal  doses  of  arsenic  trisulphid 
hydrosol  favor  it.  Small  quantities  of  the  latter  preparation  activate 
and  larger  ones  inhibit  the  uric  acid  forming  ferments  in  liver  autol- 
ysis (M.  AscoLi  and  G.  Izar  *). 

Phosphorus,  arsenic  and  antimony  inhibit  oxidation  processes.  In 
minute  doses  this  results  in  an  increased  constructive  activity;  its 
effect  may  be  compared  with  sUght  oxygen  need,  such  as  occurs  at 
high  altitudes.  In  larger  doses  the  toxic  action  comes  to  the  fore- 
ground. The  metabolism  does  not  reach  its  end  product,  weak  car- 
bonic acid,  but  there  are  formed  the  intermediary  stronger  acids 
(lactic  acid,  glycuronic  acid,  etc.);  the  difficultly  oxidizable  fats  are 
no  longer  normally  attacked;  there  is  a  fatty  degeneration  of  the 
glands  (liver,  kidneys),  subcutaneous  tissue  and  in  the  peritoneum 
and  all  the  organs  successively.  [It  is  more  probable  that  there  is 
a  change  in  the  aggregation  of  the  fat  globules  as  the  result  of  these 
poisons  (breaking  of  emulsions).  T.  Brailsford  Robertson  has 
recently  presented  this  view,  and  he  refers  to  the  fact  that  Gay  and 
Southard  observed  the  loading  of  the  gastric  epithelium  with  visible 
fat  globules  in  animals  which  have  experienced  anaphylactic  shock. 
Science  N.  S.,  Vol.  XLV,  No.  1170,  p.  568  et  seq.  Tr.]  It  is  upon 
this  very  retention  of  fat  that  the  therapeutic  employment  of  arsenic 
depends.  It  has  been  recognized  a  long  time  by  the  arsenic  eaters 
of  Steiermark  and  by  breeders.  [This  may  be  due  to  the  destruction 
of  the  protective  action  of  an  emulsostatic  substance.     Tr.] 

With  toxic  doses,  when  the  formation  of  stronger  acids  instead  of 
weak  carbonic  acid  occurs,  there  must  results  an  increased  friction  of 
the  blood  in  the  capillaries.  As  a  matter  of  fact  circulatory  disturb- 
ances are  among  the  most  characteristic  phenomena  of  phosphorus, 
arsenic,  antimony  and  lead  poisoning.  "  Generalized  dropsy  "  (edema 
resulting  from  acid  formation  in  the  tissues,  see  p.  208  et  seq.)  is  a 
symptom  of  chronic  arsenic  poisoning. 

We  must  also  regard  the  "capillary  paralysis"  due  to  arsenic 
as  caused  by  an  increase  of  the  viscosity  of  the  blood  at  the  inter- 
faces. It  must  be  specially  emphasized  that  these  statements  are 
only  working  hypotheses. 


378  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Salts. 

The  neutral  salts  of  alkalis  may  cause  injuries^  to  organs  or 
organ  groups  by  reversible  changes  in  the  condition  of  the  organ 
colloids;  strictly  speaking,  they  are  not  poisons.  We  are  unable 
to  produce  a  poisoning,  for  instance  by  the  oral  ingestion  of  moderate 
doses  of  potassium  salts,  though  this  may  be  accomplished  with 
intravenous  injections;  under  such  circumstances,  disturbances  of  the 
heart  muscle  and  the  peripheral  vessels  are  observed.  It  would  be 
worth  determining  whether  these  phenomena  are  not  to  a  great 
extent  caused  by  changes  in  the  viscosity  of  the  blood.  Hitherto, 
potassium  salts  have  not  been  purposely  employed  therapeutically 
with  this  in  view.  H.  Bechhold  and  J.  Ziegler*''  attribute  the 
favorable  action  of  a  vegetarian  diet  in  gout  to  the  generous  supply 
of  potassium  salts  which  hinders  the  precipitation  of  urates. 

The  biological  action  of  neutral  salts  has  been  studied  chiefly 
by  biologists  and  physiologists.  We  owe  to  them  valuable  contri- 
butions concerning  the  inhibition  of  irritability  (see  p.  274  et  seq.), 
the  death  of  lower  salt  and  fresh  water  organisms  in  changed  media, 
and  the  inhibition  of  the  development  of  the  eggs  of  marine 
creatures. 

It  follows  from  all  these  investigations  that  for  the  normal  function- 
ing of  the  organisms,  no  matter  whether  animal  or  plant,  high  or  low, 
a  definite  combination  of  electrolytes  is  necessary;  upon  this  the 
normal  state  of  swelling  for  the  organ  colloids  depends.  The  cations 
are  especially  important.  The  monovalent  cations  (Na,  K)  are  held 
in  check  by  small  quantities  of  divalent  ones  (Ca,  Mg) .  [See  Clowes, 
p.  38.  Tr.]  Several  examples  may  serve  to  explain  this.  For  ani- 
mal organisms  a  given  content  of  Na  ions  is  necessary,  which  may 
at  best  be  replaced  by  Li  ions.  K  ipns  are  especially  poisonous  because 
they  change  the  state  of  turgescence  of  the  organ  colloids.  Pure 
sodium  chlorid  solution  of  physiological  osmotic  pressure  behaves  as 
a  poison;  this  was  shown  by  Jacques  Loeb  on  the  fertilized  eggs 
of  fundulus  heteroclitus,  a  small  sea  anemone.  He  also  showed 
that  this  poisonous  action  was  arrested  by  the  addition  of  a  small 
amount  of  any  salt  containing  polyvalent  cations.  Substances  which 
were  themselves  very  poisonous,  such  as  barium,  zinc,  lead  and  ura- 
nium salts,  under  these  circumstances  detoxicate  sodium  chlorid,  but 
copper  and  mercury  salts  and  ferric  ions  showed  no  detoxicating 
action.  K.  G.  Lillie  *^  observed  a  similar  antitoxic  action  of  poly- 
valent cations  in  the  poisoning  of  the  larval  forms  of  arenicola,  a  sea 
annelid.     Its  ciliary  movement  is  stopped  by  pure  Na  and  Li  salts 

1  These  questions  are  treated  in  Chapter  XVII. 


TOXICOLOGY  AND  PHARMACOLOGY  379 

since  the  cilia  dissolve.  This  injurious  action  is  stopped  by  poly- 
valent cations. 

Interesting  in  this  connection  are  the  experiments  of  Wo.  Ost- 
WALD  *^  on  the  vitality  of  the  sand  flea  (gammarus  pulex)  which  lives  in 
fresh  water.  It  survives  in  sea  water  three  or  four  days  but  in  a 
mixture  of  four-fifths  sea  water  and  one-fifth  distilled  water,  it  lives 
almost  as  long  as  in  fresh  water.  If  each  constituent  of  the  sea  water 
is  successively  removed,  the  toxicity  of  the  remainder  rises,  that  is 
the  duration  of  life  diminishes  in  the  following  order: 

NaCl  +  KCl  +  CaCl2  +  MgS04  +  MgClz 

NaCl  +  KCl  +  CaCl2  +  MgSO« 

NaCl  +  KCl  +  CaCl2 

NaCl  +  KCl 

NaCl. 

According  to  W.  J.  V.  Osterhout  what  has  been  demonstrated 
for  animals  is  equally  true  for  plants  (algae,  grains,  hverwort  and 

moulds).      The  fresh  water  alga,  vaucheria  sessilis,  is  killed  in  -^^ 

NaCl  solution  but  continues  to  grow  if  a  trace  of  calcium  chlorid  ia 
added.  According  to  Chas.  B.  Lipman  the  dry  weight  of  ripe 
barley  was  increased  if  CaS04  was  added  to  a  culture  containing 
sufficient  sodium  sulphate  to  be  harmful.  In  this  case  as  with  cul- 
tures of  bacteria,  the  antagonistic  action  of  the  cations  play  an 
important  part. 

Though  we  employ  physiological  sodium  chlorid  solution  in  many 
experiments  for  the  maintenance  of  isotonicity,  it  is  merely  a  make- 
shift, and  on  this  account  there  have  recently  been  introduced  solu- 
tions which,  as  well  as  being  isotonic,  have  a  composition  similar  to 
the  blood  (Ringer's  and  Adler's  solution)  and  thus  maintain  its 
normal  state  of  swelling.     [More  recently  McClendon's.     Tr.] 

All  these  solutions  contain  the  divalent  Ca  ion.  We  have  indi- 
cated on  page  70  how  we  believe  its  detoxicating  effect  is  brought 
about;  it  opposes  the  swelling  due  to  monovalent  ions  (Na,  K). 
And  it  is  usually  assumed  that  the  "tanning"  is  hmited  to  the  plasma 
pellicle. 

Though  the  cations  are  of  major  importance  in  "balanced" 
combinations  of  salts,  the  anions  are  not  without  significance 
(J.  Loeb). 

As  was  mentioned  previously,  the  toxic  action  of  the  neutral  salts, 
is,  in  general,  reversible.  On  this  account  the  question  arises, 
whether  their  action  is  due  to  a  solution  or  an  adsorption  phenomenon 
by  the  organ  colloids.  Wo.  Ostwald  decided  the  question  in  favor 
of  the  latter  view.     In  the   adsorption   equation    (sec   p.  21)  in- 


380  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

X 

stead  of  —  (concentration  of  the  salt  in  the  dispersed  phase)  he 

placed  - ,  in  which  t  =  length  of  life;   -  is  accordingly  the  toxicity. 

1 
The  equation  becomes  -r-  =  k.   Wo.  Ostwald  experimented  with  the 

P 

sand  flea  mentioned  (gammarus  pulex)  and  with  another  small  crus- 
tacean {daphnia  magna).  He  placed  a  given  number  of  them,  e.g., 
twenty-five,  in  a  definite  quantity  of  water  (100  cc.)  of  different  salt 
concentration  and  every  two  minutes  he  observed  how  many  had 
meanwhile  died.  It  was  evident  that  the  zero  point  of  the  adsorp- 
tion curve  must  be  placed  to  coincide  with  the  normal  salt  con- 
tent of  the  organism,  and  that  either  a  dilution  or  a  concentration  of 
the  surrounding  water  is  toxic.  This  must  be  expressed  in  the  ad- 
sorption equation.     Accordingly,  the  toxicity  formula  for  neutral 

1 

salts,  when  their  concentration  is   increased,  is =  k;    in 

(c  —  n)~ 
V 
this  case  n  is  the  quantity  of  salt  normally  adsorbed  in  the  tissues. 

For  the  toxicity  of  subnormal  salt  solutions,  the  adsorption  for- 
mula becomes  -  •  C  ~  =  k.     Wo.    Ostwald  **  calls  the  latter  the 
t         p 

"formula  of  leaching."  Observed  and  calculated  results  agree  quite 
well. 

A  peculiar  place  is  occupied  by  potassium  iodid  and  iodin  com- 
pounds. With  all  of  them,  the  "iodin  action"  is  the  most  im- 
portant; we  may  even  assume  that  the  iodin  of  nonelectrolytes 
finally  becomes  an  iodin  ion.  The  emaciation  caused  by  its  pro- 
longed internal  use  and  the  atrophy  of  certain  glands  are  the  most 
characteristic  iodin  effects  upon  higher  animals.  Prolonged  use  of 
iodin  preparations,  according  to  H.  Meyer  and  R.  Gottlieb,* 
among  others,  causes  an  excessive  secretion  from  mucous  mem- 
branes, which  is  an  inflammatory  reaction.  Even  though  metab- 
olism experiments  have  not  revealed  any  constant  variations  from 
the  normal,  it  may  be  recalled  that  according  to  the  experiments  of 
H,  Bechhold  and  J.  Ziegler  (see  p.  54)  potassium  iodid  facihtates 
the  diffusion  of  a  third  substance  through  a  jelly.  All  the  phenomena 
mentioned  above  indicate  a  facilitation  of  metabolism.  As  was  to 
be  expected  potassium  iodid  (according  to  E.  Romberg)  lowers  the 
viscosity  of  the  blood,  and  according  to  O.  Müller  and  R.  Inada* 


TOXICOLOGY  AND  PHARMACOLOGY  381 

improves  its  circulation.  The  action  of  iodin  in  the  functional  dis- 
turbances of  arteriosclerosis  may  be  explained  by  this  property  since 
such  disturbances  may  be  attributed  to  a  faulty  blood  supply  to  the 
organs.  The  analysis  of  the  individual  features  of  the  process  has 
not  yet  been  completed. 

E.  Bernoulli  explains  the  action  of  bromin  salts  as  a  colloidal  ac- 
tion. Bromids,  which  are  given  as  sedatives,  induce  in  both  man  and 
beast  apathy  and  slumber  as  their  most  marked  effect.  It  may  be 
demonstrated  that  a  portion  of  the  chlorin  in  the  body  is  displaced 
by  bromin  and  that  administration  of  NaCl  induces  recovery.  E. 
Bernoulli  has  shown  that  the  brain  is  more  swollen  in  equimolecular 
solutions  of  NaBr  than  of  NaCl.  In  addition  he  was  able  to  restore 
rabbits  poisoned  with  NaBr  by  injecting,  instead  of  NaCl,  other 
salts  which  inhibit  swelling  (sodium  sulphate  and  nitrate).  Thus  it 
is  highly  probable  that  change  in  the  function  of  the  nerve  cells  in- 
duced by  bromids  may  be  attributed  to  a  swelling. 

In  the  case  of  the  alkaline  earths  there  occur  actual  specific  actions 
and  we  find  transitions  to  irreversible  conditions  which  are  induced 
by  the  salts  of  heavy  metals  on  albumin  and  lipoid  colloids.  For 
instance,  barium  has  a  very  intense  action  on  the  heart  and  the 
vascular  musculature.  Of  all  the  anions  sulphocyanid  inhibits  pre- 
cipitation least,  so  that  Wo.  Pauli  **  asserted,  a  priori,  that  a  com- 
bination of  sulphocyanid  and  barium  would  exert  an  especially 
severe  effect.  He  maintained  animals  under  the  influence  of  a 
moderate  sulphocyanid  intoxication  which,  though  the  heart  was 
strong  and  regular,  stimulated  the  vagus  and  the  vascular  centers. 
In  a  moderate-sized  dog  5  mg.  of  barium  chlorid  sufficed  to  cause  an 
immediate  stoppage  of  the  heart.  Calcium  and  strontium  salts 
acted  in  a  similar  way,  but  much  larger  doses  were  required  since 
with  these  there  is  much  less  specific  affinity  for  heart  muscle. 

C.  Neuberg  *  and  his  pupils  were  able  to  prepare  in  methyl  alco- 
hol colloidal  solutions  and  jellies  of  compounds  of  calcium,  strontium, 
barium  and  magnesium,  which  are  insoluble  in  water,  as  for  instance 
CaO,  CaS04,  CaCOs,  the  oxalate  and  phosphate  of  Ca,  ]\IgHP04, 
BaCOs,  etc.  Since  they  are  lipoid-soluble,  it  is  possible  they  are  of 
importance  in  the  animal  organism.  C.  Neuberg  believes  that  pos- 
sibly they  may  develop  in  the  cells  in  the  presence  of  sugar,  glj'cerin 
or  even  in  the  presence  of  ethyl  alcohol  in  an  aerobic  respiration;  in 
my  opinion  the  presence  of  the  body  colloids  should  suffice  to  permit 
them  to  develop.  The  blood  pressure  elevating  properties  of  barium 
salts  may  eventually  be  utilized  in  the  form  of  colloid  solutions  in- 
asmuch as  such  solutions  do  not  possess  the  undesirable  by-effects 
of  barium  salts. 


382 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Aluminium  is  the  bond  between  the  earth  alkalis  and  the  heavy 
metals.  It  coagulates  albumin  in  ''  irregular  series"  and  under  certain 
conditions  the  albumin-aluminium  precipitates  are  reversible.  In 
this  connection,  thallium  coagulates  the  protoplasm  of  aquatic  plants 
(spirogyra,  elodea,  etc.),  but  they  recover  when  replaced  in  their 
original  medium  (J.  Szücs). 

The  soluble  salts  of  heavy  metals  form  irreversible  metal  albumin 
precipitates  with  albumin  which  either  flock  out  immediately  or, 
depending  on  the  concentration  of  the  salt  solution,  persist  in  the 
colloidal  condition. 

For  this  property  of  the  salts  of  the  heavy  metals ,  besides  the 
valence,  the  electrolytic  solution  pressure  (see  H.  Bechhold  *^)  is 
determinative;  colloid  precipitation  depends  upon  these  two  factors. 
The  toxicity  threshold  of  the  various  salts  of  the  heavy  metals 
has  been  arranged  in  series.  Mathews  *  tested  it  on  the  motor 
nerves  of  frogs.  Kahlenberg  and  True,  as  well  as  F.  D.  Heald, 
tested  them  on  plant  seedlings.  I  reproduce  (from  R.  Höber)  the 
series  determined  by  Mathews  for  the  inhibition  of  the  develop- 
ment of  the  fertilized  eggs  of  the  sea  anemone,  fundulus  heteroclitus. 


Salts. 


MnCl2 

ZnCla 

CdCl2 

FeCla 

CaCl2 

NiCl2 

Pb  (CH3COO)2 

CuCl2 

HgCl2 

AgNOg 

AuCls 


Solution  pressure  in  volts. 


+0.798 
+0.493 
+0.143 
+0.063 
-0.045 
-0.049 
-0.129 
-0.606 
-1.027 
-1.048 
-1.356 


Threshold  of  toxicity. 


1/4  n 
1/800 n 
1/12,500  Ji 
1/lOn 
l/12n 
1/15  n 
1/5,000« 
1/15,000  n 
1/50,000  n 
1/90,000« 
1/20,000« 


The  exceptions  which  ZnCl2  and  CdCl2  show  (according  to  R. 
Höber)  may  depend  in  the  first  instance  upon  strong  hydrolysis 
(acid  reaction)  and  in  the  latter  on  the  smaller  amount  of  electro- 
lytic dissociation  together  with  greater  lipoid  solubility. 

For  the  antagonistic  action  of  ions  of  the  heavy  metals  see  pages 
70  and  378. 

The  intravenous  injection  of  the  salts  of  the  heavy  metals,  which 
is  associated  with  precipitation  of  protein,  causes  in  suitable  doses 
anaphylactic  phenomena  which  may  be  explained  by  what  has  been 
said  on  page  210. 

The  salts  of  the  heavy  metals  in  respect  to  their  toxicity  appear 
to  me  to  have  powerful  specific  influences.     For  instance,  copper 


TOXICOLOGY  AND  PHARMACOLOGY  383 

salts  are  powerful  poisons  to  algae,  infusoria  and  fungi.  According  to 
BoKORNY  they  are  effective  even  in  dilutions  of  1  :  100,000,000. 
Vertebrates  can  stand  them  in  relatively  higher  doses;  but  even 
among  these  there  is  considerable  variation;  cats,  for  instance,  are 
said  to  be  very  sensitive  to  copper  salts. 

Some  of  the  heavy  metal  cations  in  spite  of  always  precipitating  al- 
bumin appear  to  be  able  to  enter  the  circulation  and  to  be  definitely 
stopped  only  when  they  reach  the  filter  membranes  of  the  glands  (liver, 
spleen,  kidneys).  On  this  account  we  frequently  encounter  kidney 
irritation  from  the  toxic  heavy  metal  cations  (mercury,  lead,  etc.). 
Doubtless  their  solubilities  in  the  hpoids  are  an  important  factor. 

The  formation  of  irreversible  albumin  compounds  kills  the  cell 
which  is  involved.  [Hg,  when  absorbed  to  the  extent  of  4  mg.  per 
kilo,  slays  relentlessly.  It  forms  an  irreversible  compound,  unaffected 
by  antidotes  or  by  washing  with  water  as  has  been  shown  by  Sansum. 
Tr.]  On  this  account  besides  the  acids  and  the  alkalis,  salts  of  the 
heavy  metals,  e.g.,  copper  sulphate,  silver  nitrate  and  zinc  chlorid,  are 
used  as  caustics.  Astringents  act  by  causing  a  coagulation  of  the 
topmost  layers  of  mucous  membranes  or  inflamed  surfaces.  There- 
fore they  include  salts  of  the  heavy  metals,  as  silver  nitrate,  copper  sul- 
phate and  acetate,  zinc  sulphate  and  acetate  and  bismuth  subnitrate. 
Besides  these,  ferric  chlorid  and  the  various  aluminium  salts  (alumin- 
ium acetate,  alum,  etc.)  of  whose  powerful  flocculating  action,  resulting 
from  the  trivalence  of  Fe  and  Al  we  have  already  learned  (see  p.  84) ; 
the  flocculating  action  in  fact  depends  on  the  colloidal  ferric  hydroxid 
and  aluminium  hydroxid  contained  (see  below).  Similar  results  may 
be  obtained  with  tannin,  formaldehyd,  and  in  short  from  all  the  hard- 
ening agents  discussed  in  Chapter  XXIII,  provided  their  employment  is 
not  precluded  by  undesirable  properties  {e.g.,  picric  acid  and  osmic  acid) . 

Iron  Salts  and  Iron  Oxid  Hydrosol. 

Recent  researches  have  shown  that  only  ionizable  iron  compounds 
have  a  pharmacologic  action  (upon  the  formation  of  red  blood  cor- 
puscles in  chlorosis),  but  they  show,^  on  the  contrary,  that  prepara- 
tions with  iron  firmly  bound  (hemoglobin  preparations  in  particular) 
have  no  specific  action.  The  numerous  preparations  in  which  iron  is 
administered  as  a  colloidal  iron  oxid  {ferri  oxidat.  saccharatum  solu- 
bile, liq.  ferri  oxid.  dialys,  and  in  some  of  the  chalybeate  mineral 

^  It  may  be  mentioned  in  contradiction  to  this,  that  colloidal  Fe  (OH)  3,  ac- 
cording to  M.  AscoLi  and  G.  Izar,  favors  the  total  autolysis  of  the  liver  as  weU  as 
its  individual  factors  (see  p.  369  et  seq.)  and  that  the  ferments  taking  part  in  the 
formation  of  uric  acid  are  activated  by  the  addition  of  coUoidal  ferric  hydroxid; 
larger  quantities,  however,  inhibit  uric  acid  formation. 


384  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

waters)  are  active  only  to  the  extent  that  they  are  dissolved  in  the 
hydrochloric  acid  of  the  gastric  juice.  I  cannot  form  any  idea  as  to 
the  process  of  absorption  since  in  the  alkaline  content  of  the  small 
intestine  where  absorption  occurs,  the  iron  is  thrown  down  again 
as  a  colloidal  gel.  Those  colloidal  iron  preparations  from  which  the 
iron  ion  slowly  splits  off  (e.g.,  liquor  Jerri  albuminati,  ferratin,  etc.)  are 
preferable  since  they  exert  a  less  injurious  effect  on  stomach  and 
intestine  (indigestion  and  constipation) .  After  intravenous  or  subcu- 
taneous injection  of  iron  salts  colloidal  ferric  albuminate  compounds 
are  formed  which  may  cause  severe  anaphylactic-Hke  symptoms  of 
poisoning  (see  p.  382) .  When  iron  salts  are  taken  by  mouth  this  action 
does  not  occur,  since  the  iron  is  arrested  in  the  liver.  The  cathodal- 
migrating  positive  iron  oxid  hydrosol  precipitates  with  the  anodal- 
migrating  blood  colloids  as  an  irreversible  gel.  This  is  the  reason  why 
ferric  chlorid  is  so  suitable  for  hemostasis.  The  greater  part  of  the  Fe 
in  FeCls  exists  as  iron  oxid  hydrosol  as  the  result  of  hydrolytic  cleavage. 
When  blood  coagulates,  the  excess  of  HCl  is  bound  by  the  blood  salts. 

R.  BuNSEN,  in  his  first  scientific  paper,  showed  that  ''freshly  pre- 
cipitated ferric  hydroxid"  is  able  to  take  up  considerable  quantities 
of  arsenious  acid  and  recommended  it  on  this  account  as  an  antidote 
for  arsenic  poisoning.  W.  Biltz  *2  showed  that  the  distribution  of 
arsenious  acid  between  iron  oxid  hydrogel  and  water  has  the  charac- 
teristic of  an  adsorption  curve  and  not  that  of  a  chemical  combina- 
tion. The  protective  action  against  arsenious  acid  depends  moreover 
upon  the  method  of  preparing  the  ferric  oxid  hydrogel.  Works  on 
materia  medica  prescribe  that  it  be  freshly  prepared.  Perhaps,  the 
inhibiting  action  which,  according  to  L.  Pincussohn,*  ferric  oxid 
hydrosol  exerts  on  pepsin  digestion  depends  upon  adsorption. 

Although  colloidal  ferric  hydroxid  serves  as  the  typical  positive 
colloid  H.  W.  Fischer  *^  succeeded  in  preparing  a  negative  ferric  oxid 
hydrosol,  as  well.  He  did  this  by  pouring  ferric  chlorid  solution  into 
sodium  hydrate  solution  which  contained  glycerin  as  a  protector. 
Glycerin  and  the  excess  of  alkali  were  then  removed  by  diffusion. 
Instead  of  glycerin  other  polyvalent  alcohols,  e.g.,  mannit,  erythrit 
and  cane  sugar,  may  be  employed.  The  object  of  his  experiments 
was  to  obtain  ferric  oxid  hydrosol  which  might  be  injected  intrave- 
nously. Positive  ferric  oxid  precipitates  with  the  negative  serum 
colloids;  on  this  account  the  intravenous  injection  of  positive  ferric 
oxid  is  immediately  fatal  to  animals,  on  account  of  embolism.  A 
remarkable  exception  to  this  was  found  by  C.  Foa  and  A.  Aggazzotti  * 
in  dogs;  they  are  insensitive  to  positive  ferric  oxid;  no  explanation 
for  this  exists.  Negative  ferric  oxid  may  be  mixed  mth  serum  in  any 
proportion.     It  forms  a  deep  ruby  red  solution  which  may  at  times 


TOXICOLOGY  AND  PHARMACOLOGY  385 

take  up  much  more  than  its  own  volume  of  oxygen.  Since  it  has 
some  other  properties  of  hemoglobin  H.  W.  Fischer  calls  this  prepa- 
ration "synthetic  active  hemoglobin"  {Effectsynthese  des  Hämo- 
globins). Properly  prepared  ferric  oxid  may  be  injected  intravenously 
into  rabbits;  yet  depending  upon  how  it  was  prepared  it  proved  to 
be  more  or  less  toxic  even  though  no  embolism  could  be  discovered. 
Negative  ferric  oxid  seems  to  store  itself  up  in  the  glandular  organs 
(liver,  kidneys)  just  as  do  other  hydrophobe,  mostly  negative  col- 
loids. No  change  of  charge  occurs  since  it  is  only  after  HCl  is  added 
that  a  blue  coloration  occurs  with  potassium  f errocyanid.  Although 
positive  ferric  oxid  hydrosol  strongly  adsorbs  arsenious  acid,  its 
protective  action  is  almost  completely  lost  if  such  a  mixture  of  the 
ferric  oxid  hydrosol  and  the  adsorbed  arsenious  acid  is  injected 
subcutaneously.  Negative  ferric  oxid  hydrosol,  under  the  same 
circumstances,  exerts  a  very  considerable  protective  action,  but  fails 
completely  when  such  a  mixture  is  injected  intravenously.  H.  W. 
Fischer  attributes  this  to  the  presence  of  hemoglobin  which  tears 
the  arsenious  acid  from  the  ferric  oxid  hydrosol. 

Narcotics  and  Anesthetics. 

We  class  as  narcotics  such  substances  as  temporarily  suspend 
cerebral  function,  and  the  activity  of  the  reflex  centers.  Narcosis  is, 
therefore,  a  reversible  process. 

According  to  the  theory  of  Hans  Meyer  and  E.  Overton,  nar- 
cosis is  produced  by  such  substances  as  dissolve  especially  easily  in 
the  lipoids  of  the  plasma  pellicle  but  are  not  entirely  insoluble  in 
the  plasma.^  They  determined  the  distribution  coefficient  between 
oil  and  water  for  a  large  number  of  substances  and  found  that  those 
substances  in  which  the  distribution  coefficient  (oil  :  w^ater)  is  high 
are  good  narcotics,  e.g.,  chloroform,  ether,  acetone,  chloral  hydrate, 
urethan,  etc.  The  coincidence  is  not  only  qualitative  but  it  was 
possible  by  determining  the  "critical  concentration"  to  show  that  it 
was  quantitative.  By  "critical  concentration"  is  meant  the  con- 
centration of  a  narcotic  in  water  which  just  suffices  to  maintain  the 
narcosis  of  an  organism  (animal  or  plant).  With  over  100  substances, 
a  surprising  parallehsm  was  shown  to  exist  between  "critical  narcotic 
concentration"  and  coefficient  of  diffusion  between  oil  and  water,  so 
that  a  causative  connection  between  narcosis  and  fat  solubihty 
seems  obvious. 

^  There  exists  a  certain  parallelism  between  the  phj'^siological  action  of  nar- 
cotics and  their  abiUty  to  depress  the  surface  tension  of  water.  Upon  this  is 
based  J.  Traube's  *  theory  of  narcosis.  The  depression  of  surface  tension  favors 
the  penetration  of  the  narcotic  into  the  cell. 


386  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

In  recent  years  we  have  become  acquainted  with  a  number  of  facts 
which  cannot  be  reconciled  with  the  Meyer-Overton  theory.  For 
instance,  S.  J.  Meltzer  showed  that  magnesium  salts  possess  power- 
ful narcotic  properties.  G.  Mansfeld  and  Bosanyi  then  showed  that 
during  profound  magnesium  narcosis  there  was  absolutely  no  change 
from  the  normal  magnesium  content  of  the  brain.  No  increase  in 
Mg  was  demonstrable  either  in  the  lipoid  or  the  lipoid  free  brain 
substance.  Furthermore,  it  developed,  that  the  lipoid  solubility  of 
the  narcotics  was  to  a  certain  extent  merely  accidental  which  paral- 
leled other  physico-chemical  properties.  According  to  J.  Traube 
and  J.  Czapek  diminution  of  surface  tension  parallels  the  narcotic 
properties.  We  must  emphasize,  however,  that  in  Traube's  experi- 
ments only  the  diminution  of  the  surface  tension  to  air  was  deter- 
mined, whereas  in  the  organism  we  are  concerned  with  surface 
tensions  arising  between  two  fluids  or  between  a  fluid  and  a  gel 
phase.  The  observations  of  Battelli  and  Stern  have  less  connec- 
tion with  fat  solubility;  according  to  them  there  is  a  parallelism 
between  the  precipitation  of  certain  proteins,  the  inhibition  of  oxida- 
tions in  the  tissues  and  the  narcotizing  activity  of  narcotics.  War- 
burg and  Wiesel  showed  that  narcotics  inhibit  the  ferment  activity 
of  the  pressed  juice  of  yeast  as  well  as  of  the  yeast  cells.  Without 
discussing  the  hypothetical  basis  of  these  processes  we  may  conclude 
from  them  that  lipoid  solubility  does  not  constitute  the  sole  physico- 
chemical  basis  for  narcosis. 

At  present  the  tendency  is  to  believe  that  the  essential  factor  in 
narcosis  is  a  modification  of  the  plasma  pelhcle  which  reversibly 
changes  its  normal  permeability  for  electrolytes,  so  that  it  is  an  open 
question  whether  this  membrane  is  pure  protein  (see  p.  239  et  seq., 
membrane)  or  a  mixture  of  lipoid  and  protein  (see  also  S.  Loewe). 

An  interesting  support  for  this  view  was  supplied  by  R.  Höber 
and  his  pupil  A.  Joel  when  they  measured  the  electric  conductivity 
of  blood  corpuscles  under  the  influence  of  narcotics.  Although  it  is 
true  that  blood  corpuscles  are  not  nerve  cells  there  are  such  similari- 
ties as  justify  us  in  applying  to  nerve  cells,  observations  made  on  blood 
corpuscles.  R,.  Höber  found  that  narcotics  inhibited  the  exit  of 
electrolytes  when  dilute,  and  increased  it  when  concentrated.  Nar- 
cotics when  dilute  produce  quite  the  opposite  effect  they  do  when 
they  are  concentrated.  This  is  analogous  to  the  conductivity 
determinations  of  Osterhout  on  plant  cells  and  the  observations  of 
Sv.  Arrhenius  and  Bubanovic  as  well  as  J.  Traube  that  small 
amounts  of  many  hemolytic  agents  inhibit  homolysis. 

Obviously,  every  substance  which  dissolves  in  fat  is  not  a  narcotic; 
it  is  such  only  if  it  can  be  again  removed  from  the  lipoid  without 


TOXICOLOGY  AND  PHARMACOLOGY  387 

leaving  permanent  changes.  We  thus  arrive  at  the  chief  point  in 
the  problem.  The  Meyer-Overton  theory  explains  the  conditions 
under  which  a  substance  may  act  as  a  narcotic,  but  it  does  not  show 
why  it  narcotizes;  in  other  words,  what  the  essence  of  narcosis  is. 
Recent  investigations,  especially  those  of  R,  Höber,  have  shown 
that  narcosis  is  brought  about  by  a  change  in  the  state  of  swelling  of 
the  nerve  colloids  by  which  the  changes  which  would  otherwise  be 
induced  by  the  cell  electrolytes  upon  stimulation  are  arrested.  Ex- 
perimentally we  consider  an  organ  narcotized  if  its  irritability  is 
temporarily  arrested  or  definitely  changed.  If  we  pass  the  im- 
pulse of  an  electric  current  through  a  muscle  it  contracts.  If  the 
ends  of  a  muscle  are  attached  to  a  galvanometer  and  we  stimulate 
the  muscle  the  needle  of  the  galvanometer  makes  a  short  excursion; 
this  is  called  the  current  of  action.  This  is  associated  in  no  way 
with  the  muscular  contraction,  for  we  may  produce  an  electric  im- 
pulse in  the  nerve  the  same  way  and  nerves  do  not  contract.  The 
excursion  of  the  galvanometer  needle  is  the  only  evidence  that  the 
nerve  is  stimulated.  All  these  phenomena  are  temporarily  arrested 
as  soon  as  the  organ  is  narcotized. 

If  we  now  see  that  normal  irritability  is  manifest  as  the  result  of 
an  electrolytic  process  in  which  transitory  changes  in  turgescence 
occur,  and  that  the  turgescence  of  nerves  and  muscle  colloids  are 
changed  by  salts,  by  which  the  irritability  is  consequently  influenced, 
we  shall  not  doubt  that  there  is  a  connection  between  turgescence 
and  irritability.  When  we  find  that  the  influence  of  salts  upon  the 
swelling  capacity  of  cell  colloids,  especiaUy  the  hpoids,  is  placed  in 
abeyance  or  suspended  by  narcotics,  the  mass  of  evidence  is  con- 
clusive. 

The  connection  between  irritability  and  colloid  turgor  was  dis- 
cussed in  Chapters  XVII  and  XXI;  the  following  passages  will 
show  that  narcotics  arrest  changes  in  turgescence.  R.  Höber  *'^  has 
shown  that  the  axis  cylinders  of  nerve  fibers  swelled  up  in  some 
portions  under  the  influence  of  neutral  salts  and  shrank  in  others,  as 
is  beautifully  shown  by  staining  with  methylene  blue.  The  phe- 
nomenon is  reversible.  Swelling  under  the  influence  of  neutral  salts 
does  not  occur  when  ethyl  urethan  narcosis  is  produced  simultane- 
ously. Accordingly,  in  this  case  the  narcosis  may  be  demonstrated 
in  the  stained  sections  (see  p.  336).  A.  R.  Moore  and  H.  E.  Roaf* 
found  that  lipoid  suspensions  are  precipitated  by  small  quantities  of 
chloroform,  alcohol,  ether,  etc.,  instead  of  being  dissolved  by  them. 
R.  Goldschmidt  and  E.  Pribram  *  found  a  similar  action  of  chloral 
hydrate  and  urethan  in  lecithin  suspensions. 

According  to  S.  J.  Meltzer  magnesium  salts  produce  narcosis  if 


388 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


subcutaneously  or  intravenously  injected.     I  wish  to  call  attention 
to  the  fact  that  according  to  0.  Purges  and  E.  Neubauer  *  MgS04 

and  MgCl2  in  — r-  solution,  unlike  other  electrolytes,  have  very  nar- 
row precipitation  limits  for  lecithin  suspensions;  with  this  fact  their 
narcotic  action  possibly  stands  in  some  relation.  Lower  animals  are 
also  narcotized  by  magnesium  salts.  On  this  account  it  is  used  by 
zoologists  to  fix  objects  in  their  natural  state,  because  Mg  narcosis  is 
not  preceded  by  irritation.  There  is  still  not  very  much  evidence 
that  change  in  swelling  is  inhibited  by  narcotics;  the  evidence  must 
be  reinforced,  especially  by  simple  test-tube  experiments  on  the  rela- 
tive influence  of  salts  and  narcotics  in  changing  the  turgor  of  lipoids. 
We  see  here  a  promising  field  for  experiment.  It  may  be  possible 
to  combine  this  theory  with  that  of  Verworn's  school.  According 
to  their  view,  the  oxidation  processes  in  the  cell  are  arrested  during 
narcosis,  a  hypothesis  supported  by  numerous  experiments.  [A.  R. 
C.  Haas  has  recently  shown  that  when  Laminaria  is  exposed  to  anes- 
thetics (in  sufficient  concentration  to  produce  any  result)  there  is  an 
increase  in  respiration,  which  may  be  followed  by  a  decrease  if  the  re- 
agent is  sufficiently  toxic.  Science  N.  S.,  No.  1193,  p.  46  e^seg.  Tr.] 
In  this  connection  it  must  be  recalled,  especially,  that  oxygen  and 
carbonic  acid  are  much  more  soluble  in  lipoids  than  in  water  and  that 
narcotics  diminish  the  absorption  capacity  of  the  cell  fipoids  for  oxy- 
gen (G.  Mansfeld*).  It  would  be  interesting  to  determine  the 
extent  to  which  this  solubility  is  influenced  by  the  turgor  of  the  lipoids. 
Elsewhere  I  have  already  stated  that  the  Meyer-Overton  theory 
of  narcosis  demands  a  reversible  distribution  of  the  narcotic  be- 
tween fipoids  and  plasma.  Whether  this  distribution  occurs  as  a 
Henry's  distribution  or  as  an  adsorption  is  immaterial  in  principle 
(but  not  for  the  action!).  According  to  a  table  of  M.  Nicloux  *  the 
distribution  of  chloroform  seems  to  me  to  approach  that  of  adsorp- 
tion. After  the  termination  of  narcosis,  the  blood  of  a  dog  con- 
tained the  following  content  of  chloroform  (in  per  cent). 


Chloroform  Content  in  Per  Cent. 


After. 

First  experiment. 

Second  experiment. 

0  minutes 

0.054 

0.0255 

0.0205 

0.018 

0.0135 

0.0595 

15  minutes                   

30  minutes 

6.023 

1  hour          

0.018 

3  hours                  

0.0075 

7  hours                    

0.0015 

TOXICOLOGY  AND  PHARMACOLOGY  389 

The  action  of  narcotics  on  the  permeabihty  of  electr-olytes  is 
reversible  according  to  R.  Höber,  and  may  be  reversed  by  washing 
them  out  provided  the  amount  of  added  narcotic  is  not  too  great. 
R.  Höber  is  of  the  opinion  that  narcosis  is  characterized  by  a  change 
in  the  plasma  pellicle  in  which  the  increase  of  permeability  to  normal 
stimuli  is  inhibited. 

A  physico-chemical  study  by  S.  Loewe  actually  showed  that 
chloroform  was  adsorbed  by  the  white  matter  of  the  brain  and  that 
sulphonal,  trional  and  tetronal  were  adsorbed  by  lipoids. 

We  see  from  the  table  that,  at  first,  chloroform  disappears  very 
rapidly  but  that  the  final  portions  are  tenaciously  held.  A  similar 
table  for  ether  reveals  an  approximately  proportional  disappearance 
of  ether  from  the  blood  in  given  units  of  time,  which  would  approxi- 
mately answer  the  demands  of  Henry's  law.  The  slower  recovery 
from  chloroform  than  from  ether  narcosis  is  thus  explained. 

It  is  evident  from  what  has  been  previously  said  that  narcosis 
merely  represents  a  given  segment  of  the  curve  which  different  con- 
centrations of  the  narcotic  cause  in  the  turgor  of  the  cell  lipoids. 
The  conmiencement  of  the  curve  with  low  narcotic  concentration 
indicates  the  condition  of  irritability  before  narcosis,  the  terminal 
limb  with  high  narcotic  concentration  means  death. 

What  has  been  said  here  of  benumbing  the  entire  body  mutatis 
mutandis,  appHes,  for  the  individual  organs,  in  the  case  of  local  anes- 
thesia. Local  anesthesia  may  be  produced  by  all  sorts  of  substances  — 
by  very  dilute  caustics  (acids,  phenol),  by  distilled  water,  by  aniso- 
tonic  salt  solutions,  in  short,  by  all  substances  which  change  the 
turgor  of  the  cell  hpoids.  Practically  most  of  them  are  useless  be- 
cause the  first  portion  of  the  curve,  the  state  of  irritation  which  is 
expressed  by  pain  in  subcutaneous  injections,  is  too  prolonged;  in  the 
case  of  others,  because  the  segment  which  signifies  local  anesthesia  and 
which  hes  between  the  ''irritation  limb"  and  that  of  permanent 
damage  is  too  short;  still  in  others,  because  an  irreversible  change  in 
the  cell  colloids  may  occur  even  with  the  smallest  doses,  or  other  cell 
colloids  suffer  too  much  in  sympathy.  Practically  only  such  anes- 
thetics are  utihzable  as  produce  only  a  reversible  change  in  the  turgor 
of  the  nerve  lipoids,  as  is  exempHfied  by  cocain,  novocain  and 
anesthesin. 

It  is  not  difficult  to  range  the  other  methods  of  anesthesia,  such 
as  cold  and  the  production  of  anemia  in  this  scheme,  but  experimental 
confirmation  is  still  lacking. 

Colloid  research  also  offers  an  explanation  of  certain  by-effects  of 
narcotics.  [Evarts  A.  Graham  has  shown  that  the  toxic  action  of 
many  anesthetics  is  due  in  part  to  mineral  acids  formed  by  their 


390  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

decomposition.  He  believes  that  delayed  chloroform  poisoning 
results  entirely  from  the  destructive  action  of  HCl  formed  in  the 
tissues  and  he  attributes  the  protective  action  of  glycogen  to  the 
fact  that  the  glucose  resulting  therefrom  inhibits  the  diffusion  of 
HCl  into  gels.  The  toxic  action  of  anesthetics  has  been  shown  by 
J.  A.  Nef  to  be  due  to  an  unsaturated  carbon  atom.  The  effect  of 
such  atoms  has  not  yet  been  discussed  colloid-chemically.  (Jour. 
Amer,  Med.  Assoc,  Vol.  LXIX,  No.  20,  p.  1066  et  seq.,  quoted  by 
Graham,  loc.  cit.)  ,  . 

Bürge  attributes  the  anesthetic  action  of  anesthetics  to  the  de- 
crease in  oxidation  processes  produced  by  the  destruction  of  catalase. 
The  specific  action  on  the  nervous  system  is  due  to  the  greater  solu- 
bility of  the  lipoids  of  nervous  tissue  facilitating  the  entrance  of  the 
narcotic  into  the  nerve  cell.  Science  N.  S.,  Vol.  XLVI,  No.  1199,  p. 
618  et  seq.  Tr.]  With  large  doses  of  morphine,  chloroform  and  ether 
we  observe  more  or  less  intense  phenomena  of  irritation,  especially  in 
the  kidneys,  before  the  general  circulation  is  much  disturbed;  album- 
inuria and  hematuria  may  thus  occur.  Martin  H.  Fischer  *  (see 
p.  333)  explains  this  by  the  disturbance  in  the  oxidation  processes  of 
the  body  which  suffers  from  such  substances  and  by  the  fact  that  as 
the  result  of  the  accumulation  of  CO2  and  ultimately  of  other  acids, 
a  fixation  of  water  occurs  in  the  body  so  that  no  excess  of  water 
remains  for  excretion  by  the  kidneys.  Besides  the  anuria,  we  may 
thus  explain  the  thirst  which  such  patients  frequently  show.  Secre- 
tion of  urine  occurs  again  and  the  thirst  disappears  when  the  effect 
of  the  narcotic  wears  off,  even  though  the  patient  takes  no  water. 
Small  doses  of  ether,  alcohol,  etc.,  cause  the  reverse  phenomenon, 
since  by  increasing  the  activity  of  the  heart  they  bring  on  an  im- 
provement in  the  supply  of  oxygen.  By  this  means  not  only  a 
stronger  flow  of  blood  is  suppHed  to  the  kidneys  but  the  "free" 
filterable  water  in  the  blood  is  increased,  provided  the  oxidation 
processes  are  still  uninjured. 

Colloid  research  seems  to  me  to  have  raised  new  questions  regarding 
investigations  of  the  effects  from  the  prolonged  use  of  alcohol. 
Though  the  larger  part  of  the  alcohol  ingested  is  seized  by  the  lipoids, 
we  cannot  neglect  the  effect  upon  the  albuminous  colloids.  At 
present  we  can  only  assume  that  it  causes  a  diminution  of  swelling. 
The  extent  of  the  relationship  between  the  degenerative  changes  of  the 
cells,  arteriosclerosis,  etc.,  and  of  this  action  of  alcohol  remains  for 
future  investigations  to  determine.  [W.  Burridge  has  shown  that 
alcohol  increases  the  utilization  of  calcium  by  certain  cells.     Tr.] 


TOXICOLOGY  AND  PHARMACOLOGY  391 

Disinfection. 

By  disinfection  we  understand  the  killing  or  rendering  harmless  of 
dangerous  germs  outside  of  or  within  the  body.  Substances  which 
destroy  germs  living  on  foods,  without  being  very  harmful  to  higher 
organisms,  are  called  preservatives. 

For  simplicity  we  shall  first  consider  external  disinfection  by  chemi- 
cal means.  In  the  process  of  disinfection  a  distribution  of  the  dis- 
infectant between  the  organism  and  the  medium  first  takes  place.  This 
distribution  may  occur  either  in  the  manner  of  chemical  combina- 
tion, adsorption  or  in  accordance  with  Henry's  law.  In  the  two 
former  cases  it  is  conceivable  that  even  traces  of  the  poison  are  active, 
whereas  this  would  be  possible  in  Henry's  distribution  only  if  the 
substance  is  very  much  more  soluble  in  the  bacillus  than  in  its 
medium.  It  follows  from  the  ease  with  which  they  are  stained  that 
surface  attraction  is  of  great  importance  in  the  case  of  bacteria. 
And  in  fact  staining  and  disinfection  are  distinguished  only  by  the 
fact  that  in  the  latter  instance  the  absorbed  substance  exerts  a  par- 
ticular poisonous  action  on  the  microorganism. 

If  for  the  present  we  consider  a  microorganism  only  as  a  small 
particle  without  special  chemical  properties  and  add  to  such  a  hypo-, 
thetical  emulsion  of  bacteria,  a  dissolved  substance,  this  substance 
would  by  reason  of  the  mere  surface  attraction  have  a  tendency  to 
concentrate  on  the  surface  of  the  bacteria  to  a  greater  or  less  ex- 
tent, depending  upon  the  nature  of  the  dissolved  substance,  i.e., 
the  more  strongly  the  given  substance  diminishes  the  surface  tension 
of  the  water,  the  greater  is  the  concentration  at  the  surface.^ 
Most  bacteria  act  like  a  suspension  which  has  been  protected  by  a 
protective  colloid,  before  being  flocculated  by  neutral  salts;  they  are 
so  changed  by  boiling  or  by  agglutination  that  they  change  from 
hydrophile  to  hydrophobe  suspensions,  which  cannot  be  differentiated 
physically  from  kaolin  suspensions  or  the  like.  The  electric  charge 
is  that  of  an  inorganic  suspension,  i.e.,  negative;  it  is  discharged  by 
agglutinin.  All  these  questions  are  taken  up  in  detail  in  the  chapter 
on  "Immunity  Reactions." 

As  the  dispersed  phase,  microorganisms  are  strongly  adsorbed  by 
substances  with  great  surface  development.  (See  Fig.  52.)  Be- 
cause of  this  adsorption,  they  are  readily  held  back  in  fine-pored 
filters  such  as  Chamberland  candles  (unglazed  porcelain),  Berkefeld 
filters  (Kieselguhr),  asbestos,  wadding  or  carbon  filters. 

1  This  conception  was  originally  developed  and  established  by  H.  Bechhold 
in  the  "International  Congress  for  Apphed  Chemistry,"  London  (May  to  June, 
1909)  (see  KoUoid  Zeitschr.,  5,  22,  1909). 


392 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


Besides  the  microorganisms  directly  visible  in  the  microscope, 
there  are  others  so  small  as  to  be  microscopically  invisible,  and  only 
recognizable  by  their  pathogenic  effects.  They  are,  therefore,  called 
ultravisible.  Among  these  are  about  forty  pathogenic  germs,  among 
others,  smallpox,  rabies,  measles,  scarlet  fever  and  the  mosaic  disease 

DISINFECTING  ACTION  OF  HALOGEN   NAPHTHOLS. 
400,00(V 


350.000 


Lethal  action  on 


10,000 


Number  of  halogen  atoms. 

Co// 

— Sfaphy/ococcus 

-■> — — —     Diphf-heria 

— X — ^x— tX——     Sfrep/ococcus 
—     Pamtyphus 

Fig.  52.     (See  p.  402.) 

of  tobacco.  The  name  ultravisible  is  not  a  happy  one,  since  recently 
by  dark-field  illumination  there  have  been  recognized,  in  the  case  of 
many  infections,  minute  organisms,  which  we  are  justified  in  believing 
to  be  the  cause  of  the  diseases.  The  ultravisible  viruses  are  not  held 
back  by  ordinary  bacteria  filters;  recently  they  have  been  called 
filterable  microorganisms. 


TOXICOLOGY  AND  PHARMACOLOGY  393 

The  study  of  these  forms  of  Ufe  is  difficult  because  of  the  lack  of 
technical  methods  for  their  investigation.  Besides  dark-field  illumi- 
nation, colloid  research  has  provided  two  methods  which  have  already 
led  to  important  advances:  these  are  ultrafiltration  and  adsorption. 
By  means  of  the  Chamberland  filter  the  solution  of  virus  may  be 
freed  from  visible  bacteria.  In  order  to  concentrate  the  filterable 
germs  and  make  quantitative  tests  with  them,  they  may  be  con- 
centrated on  an  ultrafilter,  as  was  done  by  Betegh  with  hog  cholera 
virus,  Prowazek  and  Giemsa  with  variola;  or  they  may  be  adsorbed 
on  charcoal  or  clay  (as  did  Gins  with  smallpox). 

I  beheve  the  colloid  investigation  of  filterable  microorganisms  will 
yield  valuable  results,  since  they  form  a  transition  group  to  true 
colloids.  A  beginning  has  already  been  made.  Thus  Andriewsky 
has  shown  by  ultrafiltration  that  the  virus  of  chicken  cholera  is 
smaller  than  the  hemoglobin  molecule. 

It  has  been  repeatedly  observed  that  the  development  of  micro- 
organisms is  facihtated  by  the  presence  of  suspensions  or  hydrogels. 
Thus  Krzemieniewski  found  that  a  pure  culture  of  nitrifying  bac- 
teria grew  more  luxuriantly  and  bound  more  nitrogen  if  earth  or 
humus  was  added  to  the  culture  medium  and  Kasserer  found  a 
similar  effect  from  the  addition  of  colloidal  siUcates  and  phosphates 
of  iron  and  aluminum.  According  to  Ross  van  Lennep  pieces 
of  kidney,  meat,  cellulose,  etc.,  improve  the  growth  of  aneorobic 
bacteria,  yeast  and  B.  coU.  We  thus  see  that  these  microorganisms 
on  purely  physical  grounds  find  much  more  favorable  conditions  for 
growth  in  their  natural  habitat  than  in  artificial  media.  In  some 
instances  it  was  possible  to  determine  the  reason  for  this  phenomenon. 
Thus  Sohngen  *  and  also  Ross  van  Lennep  showed  that  charcoal 
and  some  other  sohds  favor  the  dissipation  of  carbonic  acid  which 
inhibits  the  growth  of  yeast.  In  other  instances  the  suspensions  or 
colloids  adsorb  oxygen  for  aerobic  bacteria,  nitrogen  for  nitrifjöng 
bacteria,  or  other  nutritive  ingredients  which  are  then  available  for 
growth  at  the  surfaces  of  the  respective  substances  (Hterature  given 
by  Sohngen  *)  .^  H.  Freundlich  *^  mentions  the  following  substances 
which  show  sHght  adsorptive  affinity:  salts  (especially  of  the  baser 
metals),  highly  dissociated  substances  (such  as  strong  acids  and  bases), 
aggregations  of  OH  groups  (sugar)  and  the  sulpho  group.     As  a 

'  Though  it  is  shown  on  page  396  that  the  distribution  of  phenol  between 
the  bodies  of  the  bacteria  and  their  environment  occurs  as  it  would  in  two  sol- 
vents, it  does  not  by  any  means  contradict  what  has  been  said  here,  since  a  dia- 
infectant  action  does  not  result  from  adsorption.  Disinfection  occurs  when  the 
disinfectant  penetrates  the  microorganism;  the  portion  which  has  penetrated 
may  very  well  comply  with  Henry's  law  (distribution). 


394:  COLLOIDS  IN  BIOLOGY  AND   MEDICINE 

matter  of  fact  but  few  disinfectants  are  furnished  by  the  inorganic 
acids  and  bases  and  by  the  salts  of  the  baser  metals.  Of  course  we 
do  not  include  such  concentrations  of  the  acids  and  bases  as  produce» 
a  direct  destruction  of  the  organized  substance.  As  a  matter  of  fact, 
substances  containing  the  phenyl  group  are  our  most  useful  dis- 
infectants, such  as  carbolic  acid,  cresol,  naphthol,  anilin  water,  etc. 
H.  Bechhold  and  P.  Ehrlich  *  by  combining  phenyl  groups 
(derivatives  of  dioxj^diphenylmethan  and  o-diphenol)  obtained  sub- 
stances of  hitherto  unequalled  disinfectant  action  (with  the  exception 
of  sublimate,  etc.)  and  even  this  action  was  greatly  increased  by  .the 
introduction  of  halogens.  The  work  of  H.  Bechhold,*^  which 
introduced  into  practice  the  halogen  derivatives  of  naphthol  and 
dicresol,  disinfectants  of  great  activity,  establishes  the  breadth  of 
this  assumption. 

A  dilute  solution  of  alkalis  or  acids  is  the  normal  environment 
for  the  majority  of  microorganisms.  Although  the  majority  of  micro- 
organisms prefer  a  more  alkaline  nutriment  corresponding  to  the 
dearth  of  H  ions  in  the  animal  organism,  there  are  other  bacteria  and 
moulds,  for  instance,  lactic  acid  bacteria,  which  require  or  prefer 
an  acid  medium,  e.g.,  the  moulds  which  grow  on  acid  fruit.  From 
this  it  follows  that  when  acids  or  alkalis  injuriously  affect  a  micro- 
organism, the  specific  vital  conditions  of  the  microorganism  in  ques- 
tion have  been  unfavorably  disturbed  and  accordingly  it  is  impos- 
sible to  speak  of  a  general  injurious  action  of  H  or  OH  ions. 

Many  salts  of  the  heavy  metals  (e.g.,  silver,  mercury  and  copper 
salts)  are  disinfectants.  Their  strong  adsorptive  power,  in  which 
sublimate  excels  all  others,  was  demonstrated  by  P.  Morawitz.* 

Adsorptive  capacity  is  only  a  condition  preliminary  to  the  exercise 
of  specific  toxic  action.  It  is  generally  accepted  in  the  case  of  salts 
of  the  heavy  metals  that  this  toxic  action  depends  on  the  forma- 
tion of  albuminates.  I  am  at  present  engaged  in  the  explanation  of 
these  phenomena  and  I  am  already  in  a  position  to  state  that  adsorp- 
tion is  by  no  means  the  most  important  factor. 

Finally,  there  are  among  the  inorganic  salts,  substances  with 
specific  activity,  e.g.,  the  fluorids,  thallium  carbonate,  sulphurous 
acid  salts,  boric  acid,  etc.  We  know  of  no  disinfectants  among  the 
sugars  or  their  related  substances  (e.g.,  glycerin).  P.  Ehrlich  and 
H.  Bechhold  *  as  well  as  H.  Bechhold  *^  have  shown  in  the  case  of  a 
large  number  of  aromatic  compounds  that  the  introduction  of  sulpho 
groups  into  a  disinfectant  considerably  diminishes  its  activity. 

Adsorption  in  water  according  to  H.  Freundlich  *^  is  favorably  influ- 
enced by  the  phenyl  group  and  the  halogens.  This  author  mentions 
as  an  example  chlorbenzoic  acid  (\  =  154),  benzoic  acid  (X  =  140). 


TOXICOLOGY  AND  PHARMACOLOGY  395 

Microorganisms. 

Microorganisms  occur  more  or  less  densely  in  their  media  as 
millions  of  minute  dots,  rods  or  threads.  They  constitute  a  dis- 
persed phase  and  as  such  obey  the  physical  laws  to  which  all  suspend 
sions  are  subject.  Collectively  they  possess  an  enormous  develop- 
ment of  surface;  and,  consequently,  surface  attraction  especially 
influences  those  substances  that  are  dissolved  by  them  (in  other  words, 
more  as  the  substance  diminishes  the  surface  tension  of  water) . 

Should  our  assumption  that  adsorption  plays  an  essential  part  in 
disinfection  be  correct,  then  the  same  substance  will  be  a  much  better 
disinfectant  in  aqueous  solution  than  when  dissolved  in  alcohol  or  in 
acetone.^  This  assumption  is  sustained  by  such  investigations  as 
have  been  undertaken.  According  to  Robert  Koch,  anthrax  spares 
were  not  destroyed  by  the  appKcation  for  100  days  of  5  per  cent  car- 
bolic acid  in  oil  nor  by  5  per  cent  carbolic  acid  in  alcohol  for  70  days, 
whereas  they  were  destroyed  after  48  hours'  exposure  to  5  per  cent 
aqueous  solution  of  carbohc  acid.  Anthrax  baccilli  were  of  undimin- 
ished virulence  after  2  days'  treatment  with  5  per  cent  carbolic  acid  in 
oil,  whereas  1  per  cent  aqueous  solution  killed  them  in  2  minutes. 
Moreover,  according  to  Reichel,*  the  distribution  of  the  phenol 
between  albumin  and  the  oil  (as  compared  with  water)  is  in  favor  of 
the  oil. 

According  to  the  researches  of  Paul  and  Krönig,  as  well  as  those 
of  Sheurlen  and  K.  Spiro,  phenol  acts  in  disinfecting  as  a  molecule 
and  not  as  an  ion.  Sodium  carbolate  which  is  strongly  dissociated 
has  a  much  weaker  action  than  phenol.  Phenol  is  less  dissociated 
in  alcohol  than  in  water,  so  that  if  it  were  merely  a  question  of 
dissociation,  phenol  should  be  a  better  disinfectant  in  alcoholic  than 
in  aqueous  solutions.  As  is  shown  by  the  following  data  taken  from 
Paul  and  Krönig's  paper,  the  facts  are  quite  the  reverse.  Anthrax 
spores  were  treated  with  the  disinfectant,  according  to  the  marble 
method,  and  then  sown  on  agar;  the  resulting  colonies  were  counted. 

Number  of 
colonies. 

4  per  cent  carbolic  acid  in  water 1505 

4  per  cent  carbolic  acid  in  alcohol oo 

We  thus  see  that  in  disinfection  adsorbability  from  water  is  more 
important  than  solubility. 

1  In  disinfecting  the  hands  and  skin,  alcohol  and  alcoholic  solutions  and  even 
acetone  are  almost  exclusively  used,  though  entirely  different  factors  are  of  im- 
portance in  determining  their  use  (better  capacity  to  wet  the  fatty  epidermis,  the 
shrinking  action  of  alcohol  and  deeper  penetration  into  the  capillary  spaces  of  the 
skin  (Bechhold)). 


396  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Cresol  is  less  soluble  than  phenol  and  is  a  stronger  disinfectant  than 
the  latter.  Its  solubility  in  water  is  so  limited  that  it  must  be  dis- 
solved with  the  aid  of  soaps  and  similar  substances.  These  are  not 
true  solutions;  they  are  manifest  emulsions  in  the  dark  field  (Frei 
and  Margadant),  It  is  still  an  open  question  whether  the  effect  on 
bacteria  is  exerted  by  an  envelopment  by  the  individual  cresol  soap 
droplets,  thus  forming  about  them  a  highly  concentrated  disinfectant 
film.  Another  possibihty  is  that  the  bacteria  withdraw  dissolved 
cresol  from  their  environment,  and  that  cresol  diffuses  from  the 
droplets  to  an  equal  extent  into  the  water. 

A  group  of  disinfectants  are  active  even  in  a  dilution  in  which  the 
substance  is  no  longer  chemically  demonstrable.  According  to  R. 
Koch,  interference  with  the  growth  of  anthrax  bacilli  is  caused  by 
sublimate  even  in  a  dilution  of  1  :  600,000.  According  to  H.  Bech- 
HOLD  and  P.  Ehrlich*  tetrachlor-o-diphenol  interferes  with  the 
growth  of  diphtheria  bacilli  in  a  dilution  of  1  :  400,000  to  1  :  640,000. 
According  to  H.  Bechhold,*^  tribrom-naphthol  inhibits  the  growth 
of  staphylococci  in  a  dilution  of  1  :  250,000.  We  can  understand  the 
effect  of  such  traces  of  substances  if  we  consider  the  course  of  the 
adsorption  curve  (see  p.  20)  in  which  the  distribution  between  ad- 
sorbent and  solvent  occurs  in  such  a  way  that  the  dissolved  sub- 
stance is  practically  completely  adsorbed  in  the  weaker  concentra- 
tion, whereas  in  higher  concentrations  the  distribution  approaches 
that  required  by  Henry's  law  (between  two  solvents). 

The  objection  may  be  raised  that  the  same  conditions  are  fulfilled 
in  a  purely  chemical  combination,  to  which  we  may  reply  that  in 
many  instances  such  a  chemical  combination  must  be  considered  to 
occur. 

In  favor  of  adsorption,  there  are  two  distinct  phenomena,  inhibition 
and  death.  By  choosing  a  suitable  disinfectant  in  sufficient  concen- 
tration and  exposure,  microorganisms  may  be  completely  killed;  that 
is,  they  cannot  under  any  circumstances  be  brought  back  to  life.  In 
other  cases,  it  is  only  necessary  to  remove  the  disinfectant,  to  dilute  it 
or  to  transfer  the  germs  to  another  environment,  for  the  germs  to  start 
multiplying  again;  such  action  is  called  inhibition.  In  such  a  case, 
we  must  assume  that  the  reaction  between  microorganism  and  dis- 
infectant is  reversible.     In  killing,  the  process  may  be  irreversible.^ 

1  I  can  readily  imagine  that  death  may  occur  in  a  reversible  process  if  the 
action  of  the  disinfectant  persists  for  a  sufficient  length  of  time  to  nullify  other 
vital  processes.  To  give  a  very  crude  comparison,  if  a  man  is  drowned,  the 
water  cannot  be  regarded  as  a  poison  though  it  depresses  necessary  vital  proc- 
esses. A  man  who  cannot  be  resuscitated  after  a  submersion  lasting  5  minutes 
has  fixed  no  more  water  in  his  body  than  one  who  has  been  resuscitated  after  2 
minutes'  submersion. 


TOXICOLOGY  AND  PHARMACOLOGY  397 

If  the  disinfectant  were  a  firm  combination  with  the  microorganism 
it  would  be  difficult  to  explain  how  the  germ  could  multiply  again 
when  removed  from  the  disinfectant  solution.  This  is  readily  under- 
stood if  we  assume  that  the  union  between  microorganism  and  dis- 
infectant is  an  adsorption.  In  that  case  the  disinfectant  will  pass 
into  the  absolutely  indifferent  solvent  so  that  the  microorganism 
having  become  free  again  (from  the  disinfectant)  is  in  a  condition  to 
continue  its  development. 

A  few  examples  will  explain  the  foregoing.  R.  Koch  performed 
certain  experiments  in  the  following  way:  he  dried  germs  on  silk 
threads  and  subjected  them  for  a  given  time  to  a  disinfectant  solu- 
tion; after  this  he  placed  them  in  nutrient  bouillon  or  in  gelatin; 
if  the  germs  developed,  he  considered  that  the  disinfectant  was 
active;  if  they  did  not,  that  it  was  inactive.  In  this  way  R.  Koch 
subjected  anthrax  spores  for  two  days  to  5  per  cent  carbohc  acid  and 
found  that  afterwards  they  did  not  develop  in  gelatin.  B.  Riedel, 
in  the  Imperial  Health  Office,  found  that,  even  after  14  days  of  im- 
mersion in  5  per  cent  carbolic  acid,  the  germination  of  anthrax  spores 
was  not  inhibited  if  the  silk  threads  were  first  washed  with  water 
and  then  placed  in  fluid  gelatin;  the  gelatin  and  silk  threads  were 
thoroughly  mixed  by  prolonged  agitation  of  the  test  tube. 

According  to  R.  Koch,  a  single  immersion  of  anthrax  spores  in 
1  :  5000  sublimate  solution  suffices  to  destroy  them.  J.  Geppekt  * 
found  that  the  same  concentration  acting  four  seconds  longer,  on 
one  trial,  produced  their  death  and  on  another  did  not.  Among 
eountless  experiments  on  this  point  we  shall  mention  those  of  Eisen- 
BERG  and  Okolska  because  of  the  method  they  employed. 

They  mixed  uniform  quantities  of  disinfectant  and  bacteria,  some- 
times adding  the  entire  quantity  of  bacteria  at  once,  and  sometimes 
in  fractions.  If  the  phenomenon  is  reversible,  the  results  in  both 
cases  should  be  the  same;  if  it  is  irreversible  there  should  be  a  point 
in  the  fractioning  experiment  when  the  disinfection  should  prove  less 
satisfactory.  As  was  to  be  expected  from  other  considerations,  the 
action  of  phenol  proved  to  be  reversible  and  that  of  KMn04  and 
HgCU  to  be  partly  irreversible  (in  these  instances  the  time  of  action 
was  an  important  factor). 

Numerous  experiments  have  been  performed  in  an  attempt  to  test 
quantitatively  the  views  given  here;  the  results  actually  satisfy  the 
hypothesis  in  some  instances.  An  exact  agreement  between  obser- 
vation and  calculation  is  not  to  be  expected  because  in  disinfection 
adsorption  is  not  the  only  factor,  though  it  is  chiefly  accountable 
for  the  action  of  the  disinfectant  on  the  microorganism  (lipoid  solu- 
bility, modification  of  protoplasm,  etc.). 


398  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

The  question  of  adsorption  may  be  solved  m  one  of  two  ways 
which  I  shall  call  respectively  the  chemical  and  the  hiological 
methods. 

The  chemical  method  regards  the  microorganisms  as  a  lifeless  sus- 
pension. Suspensions  are  shaken  with  various  laiown  dilutions  of 
the  disinfectant,  and  after  the  suspension  is  removed  the  amount  of 
disinfectant  remaining  in  the  fluid  is  chemically  determined.  From 
this  we  learn  how  much  has  been  absorbed  by  the  microorganism  in 
the  various  dilutions.  It  is  the  same  method  that  is  usually  emplo^^ed 
in  chemical  adsorption  experiments.  It  may  be  criticized  because  it 
determines  the  amount  of  disinfectant  absorbed  by  the  microorganism 
but  not  the  result  of  the  adsorptive  action,  the  disinfection.  From 
a  concentrated  solution  much  more  disinfectant  is  removed  than  is 
necessary  for  killing  or  inhibition. 

R.  0.  Herzog  and  Betzel  *  employed  the  chemical  method,  with 
yeast  as  the  microorganism.  They  obtained  an  adsorption  curve 
for  chloroform  and  silver  nitrate  and  a  chemical  combination  for 
formaldehyd.  The  results  are  interesting  inasmuch  as  chloroform 
obviously  acts  by  reason  of  its  lipoid  solubility;  I  question  whether 
the  precipitation  of  albumin  by  silver  nitrate  is  the  only  factor  which 
deterixdnes  its  disinfectant  action.  The  result  for  formaldehyd  is 
especially  surprising;  its  powerful  inhibitive  action  on  development 
is  well  known,  however  its  lethal  action  was  discovered  to  be  much 
weaker.  We  shall  await  with  great  interest  the  further  prosecu- 
tion of  Herzog's  experiments  which  promise  an  explanation  of  some 
of  the  questions  proposed.  The  results  with  -phenol  are  quite  compli- 
cated. According  to  Reichel,*  in  an  aqueous  solution  of  phenol 
there  is  a  distribution  in  accordance  with  Henry's  law,  i.e.,  as  if  it 
were  distributed  between  two  solvents.  This  was  demonstrated  by 
Reichel  *  in  the  distribution  of  phenol  between  water  and  oil,  albu- 
min, Cholesterin  and  the  bodies  of  bacteria.  This  explains  why  phenol 
is  active  only  in  relatively  high  concentration.  Increasing  NaCl  con- 
tent shifts  the  relative  distribution  in  the  direction  of  the  nonaqueous 
phase.  According  to  Reichel  the  disinfectant  action  depends  on 
the  fact  that  phenol  causes  a  shrinking  of  the  albumin  phase;  this  is 
strengthened  by  the  NaCl.  In  this  way,  the  views  developed  by 
K.  Spiro  and  J.  Bruns  *  are  revived  in  modified  form. 

R.  O.  Herzog  and  Betzel  obtained  an  adsorption  curve  on  treat- 
ing yeast  with  a  phenol  solution  weaker  than  one  per  cent.  These 
contradictory  results  may  probably  be  explained  by  the  primary 
a(isorption  of  the  phenol  at  the  surface  of  the  bacterial  cell  which 
then  in  some  way  absorbs  it  until  the  body  of  the  bacterium  is 
filled.     This  I  infer  from  the  experiments  of  E.  Küster  and  Rothatjb 


TOXICOLOGY  AND  PHARMACOLOGY  399 

who  show  that  upon  the  death  of  the  bacteria  a  part  of  the  phenol 
is  Kberated. 

The  biological  method  regards  the  rapidity  of  death  (measured  by 
the  number  of  surviving  bacteria)  in  known  concentrations  of  the 
disinfectant  and  during  a  known  time  for  action.  In  this  case,  the 
changes  in  concentration  by  means  of  the  adsorbing  microorganism 
are  not  considered,  as  in  the  chemical  method,  but  only  the  damage 
to  the  microorganism.  The  method  assumes  that  "the  rate  at  which 
the  solution  of  a  substance  acts  as  a  disinfectant  is  proportional  to  the 
amount  adsorbed  from  this  solution"  (Morawitz*).  This  method 
also  is  open  to  the  objection  that  microorganisms  are  noi  a  single 
mass  with  uniform  vitality  but  a  mixture  in  various  stages  of  growth 
and  with  varying  resistance;  so  that  it  is  possible  that  the  curves 
obtained  do  not  represent  the  course  of  an  adsorption  in  various 
concentrations,  but  express  the  resistance  at  various  stages  of 
growth. 

These  criticisms  are  offered  to  show  the  difficulties  encountered 
in  an  experimental  test. 

We  may  count  in  this  group,  also,  the  experiments  in  which  an 
insight  into  the  mechanism  of  disinfection  may  be  obtained,  by 
varying  the  number  of  bacteria  with  known  changes  in  the  concen- 
tration of  the  disinfectant  acting  for  a  constant  time  (Eisenberg, 
Okolska)  . 

As  a  result  of  biological  methods,  Paul,  Birstein  and  Reuss  *  came 
to  the  conclusion  that  the  death  of  dried  adherent  staphylococci  in 
oxygen  or  in  mixtures  of  oxygen  and  nitrogen  is  due  to  the  adsorption 
of  oxygen  by  the  cocci. 

P.  MoRAWiTz^  (loc.  cit.)  found  a  good  agreement  between  the 
figures  obtained  by  Krönig  and  Paul,  upon  kilhng  anthrax  spores 
with  sublimate  and  the  formula  for  adsorption. 

Accordingly,  we  learn  from  the  quantitative  tests  that  the  dis- 
tribution of  a  disinfectant  between  microorganism  and  solution 
may  possess  the  formula  of  a  chemical  combination  (formaldehyd)  of 
adsorption  (chloroform,  silver  nitrate)  and  of  distribution  in  solvents 
in  accordance  with  Henry's  law  (phenol).  In  the  following  pages 
we  shall  see  that  transitions  between  these  different  kinds  of  distri- 
bution occur. 

It  would  certainly  be  an  error  to  regard  distribution  as  the  essential 
factor  in  disinfection.  As  a  result  of  adsorption  the  germ  is  sur- 
rounded by  a  highly  concentrated  film  of  disinfectant  whose  action 

^  This  calculation  is  referred  to  in  the  communication  of  H.  Freundlich  which 
is  mentioned  in  the  paper  of  H.  Bechhold  *  on  Disinfection  and  Colloid  Chem- 
istry, page  23. 


400 


COLLOIDS  IN  BIOLOGY  AND  MEDICINE 


destroys  it  much  sooner  than  would  be  expected  from  the  extremely- 
dilute  solutions  employed;  the  germ  retains  the  disinfectant  in  other 
media  or  in  an  infected  organism  and  only  subsequently  succumbs 
to  the  damage  the  disinfectant  inflicts.  We  must  seek  the  essential 
activity  of  the  disinfectant  in  a  modification  of  the  living  substance 
with  which  the  disinfectant  combines  or  changes  so  that  its  vital 
function  is  suspended. 

I  know  of  no  experimental  investigations  which  show  what  part  of 
the  disinfectant  is  combined  (fixed)  and  what  part  is  adsorbed,  though 
such  studies  are  very  desirable  as  they  would  afford  us  clearer  in- 
sight into  the  nature  of  disinfection,  and  they  would  also  be  of  great 
practical  significance.  For  the  present  we  must  be  satisfied  with 
analogies  which  without  question  can  be  applied  correctly  to  the 
principle  of  disinfection.  Chemically,  the  microorganisms  have  so 
much  similarity  to  textile  fibers,  especially  with  wool  and  silk  (to 
mention  only  the  great  similarity  in  staining),  that  we  may  properly 
employ  in  argument  the  results  of  W.  Schellens.  He  shook  1  gm. 
of  fiber  with  50  cc.  of  a  subhmate  solution  containing  1  per  cent  Hg 
and  found : 


Hg  fixed. 

Hg  ad- 
sorbed.' 

Hg  re- 
moved 
from  the 
solution. 

From  sublimate  (containing  1%  Hg) : 
Fruit  hairs  of  eriodendron 

Per  cent. 
1.20 

1.69 
1.9 

5.89 

indeterminable  trace 

u 

li 

0.5 

6.5 

5.2 

9.8 

12.3 

Per  cent. 

3.91 

3.08 

4.14 

12.36 

3.14 
3.0 
3.5 
4.55 

8 
6.8 

7.7 
8.2 

Per  cent. 

5.11 

Jute 

4.77 

Silk 

6.04 

Wool 

18.25 

From  mercuric  cyanid   (containing   1% 
Hg): 
Fruit  hairs  of  eriodendron 

3.14 

Jute 

3.0 

Silk 

3.5 

Wool 

5.05 

From  mercuric  acetate   (containing  1% 
Hg): 
Fruit  hairs  of  eriodendron 

14.5 

Jute 

12.0 

Silk 

17.5 

Wool 

20.5 

1  The  adsorption  figures  were  calculated  by  me  from  the  figures  of  Schellens. 

These  figures  are  interesting  from  various  points  of  view.  We 
see  that  in  the  case  of  sublimate,  of  3  parts  of  Hg,  approximately 
2  parts  are  adsorbed  and  only  one  is  fixed.  Mercuric  cyanid  is  the  one 
substance  which  is  only  adsorbed  and  suffers  practically  no  fixation ; 
though  it  is  true  it  powerfully  inhibits  development,  it  has  but  a 


TOXICOLOGY  AND  PHARMACOLOGY 


401 


weak  destructive  action.  According  to  K.  Spiro  and  J.  Bruns,*  as 
well  as  Paul  and  Krönig,  the  figures  show  that  mercuric  cyanid  is 
far  inferior  to  sublimate  as  a  disinfectant. 

We  see  from  the  table,  moreover,  in  the  case  of  mercuric  acetate, 
which  is  more  strongly  fixed  and  more  strongly  adsorbed  than  HgCl2, 
that  fixation  and  adsorption  are  not  in  themselves  alone  sufficient  for 
strong  disinfection;  the  disinfectant  must  be  offered  in  a  suitable 
form.  Mercuric  actetate  is  less  ionized  than  HgCl2,  and  since, 
according  to  Paul  and  Krönig,  as  well  as  Scheurlen  and  Spiro,  the 
Hg  ion  is  responsible  for  the  disinfectant  action,  mercuric  acetate  is 
weaker  than  sublimate. 

An  especially  convincing  proof  of  the  specific  chemical  action  of  the 
disinfectant  on  the  hving  substance  seems  to  me  to  be  that  there  is  a 
difference  in  the  resistance  of  various  groups  of  bacteria  to  disinfect- 
ants. Whereas  anthrax  spores,  tubercle  bacilli,  etc.,  show  an  enor- 
mous resistance,  cholera  vibrios,  gonococci  and  streptococci  succumb 
to  even  sHght  chemical  attacks.  The  other  groups  of  bacteria  are 
ranged  between  these  two  extremes  —  typhoid,  B.  coli,  staphylo- 
cocci, diptheria  bacilli,  etc. 

Were  merely  the  strength  of  adsorption  responsible  for  the  disin- 
fectant action,  we  could  readily  understand  that  substances  of  differ- 
ent disinfectant  power  would  exist;  we  would  understand  for  instance 
that  cresol  has  a  stronger  action  than  naphthol,  but  in  that  case  cresol 
would  always  possess  a  stronger  action  than  naphthol,  both  on  B.  coli 
and  on  typhoid  bacilli,  as  well  as  on  streptococci.  If  we  found,  how- 
ever, that  lysol  was  more  active  against  one  microorganism  and  that 
/3-naphthol  was  more  active  against  others,  we  could  attribute  the 
action  to  general  physical  properties  among  which  we  might  include 
adsorption,  but  we  would  then  have  to  ascribe  it  to  the  difference  in 
behavior  caused  by  specific  inherent  chemical  differences  in  the  bac- 
teria affected.  This  might  be  either  a  variation  in  the  solubility 
of  the  bacterial  pellicle  or  a  variation  in  the  grouping  of  the  atoms 
in  the  body  of  the  bacteria  so  as  to  manifest  a  greater  or  less 
affinity  to  the  disinfectant;  in  either  case  the  important  factor  is 
the  chemical  difference  in  the  microorganism.  Such  cases  actually 
exist  as  has  been  demonstrated  by  H.  Bechhold.*^  He  showed 
that  the  minimal  lethal  dose  in  24  hours  is: 


For  lysol  (the  cresol  content  being  com- 
pared)   

For  /3-naphthol 


Diphtheria  bacilli. 


1  :  20,000 
1  :  10,000 


B.  coli. 


1  :800 
1  :  8000 


402  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

In  accordance  with  this,  lysol  acts  twice  as  powerfully  against 
diphtheria  bacilli  as  does  (8-naphthol,  whereas  it  has  only  one-tenth 
the  effect  of  the  latter  on  B.  coli.  He  showed  further  that  a  mixture 
of  tri-  and  tetrabrom-/3-naphthol  in  one  per  cent  solution  killed  staphy- 
lococci in  from  two  to  three  minutes,  whereas  lysol  dilutions  containing 
one  per  cent  cresol  took  more  than  ten  minutes  to  do  so.  Conversely, 
a  5  per  cent  lysol  solution  containing  2.5  per  cent  cresol  is  lethal 
for  tubercle  bacilli  within  four  and  a  half  hours,  whereas  a  solution 
of  tri-  and  tetrabrom-/3-naphthol  of  corresponding  strength  had  no 
effect  even  at  the  end  of  twenty-four  hours.  We  see,  therefore, 
that  tri-  and  tetrabrom-/3-naphthol  surpass  cresol  in  its  action  upon 
streptococci,  while  upon  tubercle  bacilli  the  cresol  acts  more  power- 
fully. H.  Bechhold  *^  examined  naphthols  containing  1,  2,  3  or  more 
bromin  or  chlorin  atoms  with  reference  to  their  effect  on  various 
bacteria.  He  found,  that  with  the  admission  of  the  halogens  the 
effect  upon  various  bacteria  sometimes  increased,  that  at  times  it 
decreased,  and  that  certain  optima  could  be  obtained  (see  Fig.  52  on 
page  392).  Thus  the  maximum  disinfectant  action  against  staphy- 
lococci is  obtained  with  tri-  and  tetrabrom-;8-naphthoV  while  for  B. 
paratyphoid  it  is  obtained  with  dibrom-jS-naphthol  and  so  on.  Eisen- 
BERG  has  recently  determined  partly  specific  activities  for  a  large 
number  of  coal-tar  dyes. 

It  follows  from  this  that  to  test  an  antiseptic  on  only  one  kind  of 
bacteria  is  an  absolutely  inadequate  method  for  testing  disinfectants; 
it  is  necessary  to  subject  a  number  of  different  types  of  bacteria  to 
investigation. 

The  presence  of  a  third  substance  is  a  factor  in  the  action  of  a 
disinfectant  that  cannot  be  neglected.  We  have  already  called  at- 
tention on  page  383  to  the  influence  of  the  solvent.  To  Paul  and 
Krönig,  as  well  as  to  Scheurlen  and  Spiro,  belongs  the  credit  of 
having  made  clear  the  significance  of  electrolytic  dissociation  for 
disinfectant  action.  Dissociation  may  be  increased  or  diminished 
by  adding  certain  substances  to  the  disinfectants.  The  ionization 
of  HgCl2  is  decreased  by  the  addition  of  NaCl,  and  since  it  is  the  Hg 
ion  which  is  of  importance  in  disinfection,  the  addition  of  common 
salt  diminishes  the  disinfectant  action  of  sublimate.  On  the  other 
hand,  the  disinfectant  action  of  carbohc  acid,  cresol  and  the  other 
phenols  is  decidedly  increased  by  common  salt.  Since  NaCl  can 
have  no  effect  on  the  electrolytic  dissociation  of  phenols,  we  must 
seek  some  other  explanation.     Again,  the  nearest  comparison  must 

1  Tribrom-;8-naphthol  is  sold  under  the  trade  name  "  providoform  "  by  the  Pro- 
vidogesellschaft  (Berlin)  and  has  proven  useful  in  connection  with  the  pus  cocci 
and  diphtheria  bacilli. 


TOXICOLOGY  AND  PHARMACOLOGY  403 

be  drawn  from  the  process  of  dyeing:  common  salt  or  sodium  sul- 
phate is  frequently  added  in  dyeing  cotton  in  order  to  get  a  more 
rapid  and  complete  utilization  of  the  bath.  The  simplest  explana- 
tion of  this  is  that  there  has  been  diminution  in  the  solubility  of  the 
dye  by  means  of  the  added  salt  (i.e.,  the  dye  is  made  more  colloidal) 
and  as  a  result  of  this  a  stimulation  of  adsorption  occurs.  This  idea 
guided  Spiro  and  Bruns  *  in  their  experiments.  They  found  that 
salts  and  other  substances  which  did  not  ''salt  out"  phenol  from 
aqueous  solutions,  such  as  sodium  benzoate,  urea,  glycerin,  etc.,  had 
no  effect  in  strengthening  the  disinfectant  action  of  phenol.  Pyro- 
catechin  may  be  precipitated  by  ammonium  sulphate  but  not  by 
common  salt;  the  former  increases  the  disinfectant  action  of  pyro- 
catechin,  while  the  latter  does  not.  It  is  also  interesting  that 
according  to  Paul  and  Krönig  equimolecular  quantities  of  salts 
added  to  a  4  per  cent  carbolic  solution  increases  its  action  in  the  fol- 
lowing order:  NaCl  >  KCl  >  NaBr  >  Nal  >  NaNOs  >  CsHgONa. 
According  to  Spiro  and  Bruns,*  the  same  order  obtains  for  the 
precipitating  action  of  these  salts  on  phenol;  however,  the  sulphates 
exert  a  much  more  powerful  effect.  The  close  relationship  of  this 
series  of  salts  to  albumin  precipitation  and  to  many  other  biological 
processes  is  quite  obvious  (see  pp.  81  and  272).  Frei  and  Marga- 
DANT  have  determined  similar  relations  between  both  the  increased 
activity  of  cresol  soap  solutions  by  salts  of  the  hght  and  of  the  heavy 
metals  as  well  as  the  decreased  surface  tension  induced  by  such  salts. 

We  may  imagine  that  there  is  yet  another  possible  way  for  salts 
or  other  substances  to  exert  an  influence  by  their  mere  presence. 
H.  Bechhold  and  Ziegler*-  showed  that  the  permeability  of  jellies 
was  influenced  by  certain  substances,  and  from  this  we  may  assume 
that  the  permeability  of  the  bacterial  plasma  pellicle  for  a  disin- 
fectant may  be  changed  by  the  presence  of  a  third  substance. 

This  assumption  is  reinforced  by  experiments  of  Eisenberg  and 
Okolska  which  showed  that  alcohol,  alkalis,  urea  and  some  other 
substances,  which  increase  the  permeabiHty  of  jellies,  also  increase 
the  disinfectant  activity  of  many  antiseptics. 

In  practice  the  conditions  are  compHcated  enormously.  We  are 
no  longer  concerned  with  the  distribution  of  the  disinfectant  between 
solvent  and  microorganisms  but  organic  substances  are  added 
(sputum,  albmnin,  feces)  so  that  we  have  the  sums  of  unknown 
factors  which  can  only  occasionally  be  resolved.  The  action  of  a 
disinfectant  is  usually  much  depressed  by  organic  matter.  This  is 
also  the  reason  why  disinfection  of  the  organism,  an  internal  disinfec- 
tion or  antisepsis,  has  so  seldom  been  accomplished  by  chemical 
means.     There  are,  indeed,  substances  so  slightly  toxic,  that  men  or 


404  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

animals  may  take  the  dose  theoretically  necessary  to  disinfect 
the  body,  for  instance,  tetrabrom-o-cresol  and  hexabromdioxydi- 
phenylcarbinol  which,  according  to  H.  Bechhold  and  Ehrlich, 
stop  the  development  of  diphtheria  bacilli  in  bouillon  at  a  dilution 
of  1  :  200,000.  In  the  organism  they  have  no  effect  at  all,  in  spite  of 
the  fact  that  there  may  be  introduced  into  the  body  without  harm, 
doses  which  are  one  hundred  times  that  necessary  to  inhibit  the 
development  of  the  bacteria  in  vitro  or  to  kill  them  within  twenty- 
four  hours.  Tetrachlor-o-diphenol  behaves  similarly;  it  inhibits 
development  of  diphtheria  bacilli  in  dilutions  of  1  :  400,000  to 
1  :  640,000.  Individual  colonies  still  grew  in  a  serum  culture  in  the 
presence  of  the  chemical  at  a  dilution  of  1  :  10,000.  We  might 
question  whether  the  result  was  due  to  favorable  vital  conditions  in 
serum  removed  from  a  living  organism  or  to  other  causes.  Experi- 
ment proved  the  latter  view  correct.  By  ultrafiltration  the  free 
tetrachlor-o-diphenol  was  separated  from  the  fraction  bound  to 
serum  colloids  and  it  was  found  that  87.5  per  cent  of  the  disinfectant 
had  been  fixed  by  the  serum  colloids. 

The  relatively  simple  conditions  in  the  disinfection  of  skin  and  hands 
are  especially  instructive.  The  hands  adsorb  soHd  particles  from 
the  air  and  particles  of  dirt  and  bacteria  from  dirty  water  (H.  Bech- 
hold). Upon  washing  with  soap  these  particles  are  surrounded  by 
fatty  acids  or  fatty  acid  alkali  hydrolytically  split  off  and  cease  to 
cling  to  the  hands.  A  priori  we  might  conclude  that  there  would  be 
a  diminution  in  germs  or  disinfection  associated  with  the  cleaning  of 
the  hands;  indeed,  it  was  shown  by  earher  investigators  and  recently 
by  H.  Reichenbach  that  soaps  possess  considerable  germ-killing  action. 
I  was  able  to  prove  that  there  exists  absolute  parallelism  between 
the  detergent  and  the  disinfecting  action  of  soaps.  It  is  impos- 
sible to  disinfect  the  hands  with  soap  in  any  practicable  time 
(10  minutes),  though  this  can  be  readily  accomplished  with  alcohol 
and  alcohohc  solutions.  According  to  H.  Bechhold,  the  reason  is 
that  alcohol  with  its  low  dynamic  surface  tension  readily  enters  the 
capillary  interspaces  of  the  bare  hand  where  the  bacteria  are  lodged, 
but  aqueous  solutions,  on  the  contrary,  enter  them  very  slowly. 
This  can  be  readily  discovered  by  the  difference  in  the  distance  they 
ascend  in  strips  of  filter  paper. 

[As  the  result  of  trench  warfare  the  study  of  antiseptics  in  the 
treatment  of  wounds  has  received  intensive  study.  Antiseptic  sur- 
gery has  been  revived.  Carrel  and  Dehelly  have  elaborated  a 
valuable  system  for  the  treatment  of  wounds  by  irrigation  with  anti- 
septics of  the  chlorin  group.  The  whole  subject  of  ;wound  irrigation 
has  been  restudied  and  new  antiseptics  discovered. 


TOXICOLOGY  AND  PHARMACOLOGY  405 

J.  F.  McClendon,  in  the  Journal  of  Laboratory  and  Clinical  Medi- 
cine, August,  1917,  discusses  "The  Relation  of  Physical  Chemistry 
to  the  Irrigation  of  Wounds."  He  emphasizes  the  importance  of 
protecting  the  tissues  from  the  effects  of  prolonged  diffusion.  The 
action  of  the  antiseptics  employed  is  oxidative.  "Oxidizing  sub- 
stances are,  however,  reduced  by  cells  and  an  ideal  local  antiseptic 
would  be  one  whose  reduction  product  is  indifferent.  Hydrogen 
peroxide  falls  in  this  class  but  is  not  a  powerful  oxidizing  agent 
and  is  decomposed  by  catalase  so  rapidly  as  to  render  a  large  per- 
centage of  it  ineffective.  It  acts  as  a  mechanical  cleanser.  If 
infusoria  are  placed  in  a  solution  of  H2O2,  the  latter  penetrates  their 
protoplasm  and  is  decomposed  on  the  inside  with  the  Hberation  of 
bubbles  of  oxygen  which  burst  and  destroy  the  cells.  More  useful 
agents  are  iodin  and  chlorin,  especially  the  latter  since  HCl  formed 
on  its  reduction  may  be  neutrahzed  by  NaHCOs  that  has  been  added, 
and  thus  rendered  indifferent.  According  to  Dakin  and  his  col- 
laborators, chlorin  forms  chloramines  when  it  acts  on  protoplasm,  and 
these  chloramines  have  an  antiseptic  action.  It  is  true,  however, 
that  chlorin  oxidizes  many  organic  compounds  with  the  liberation 
of  HCl.  Chlorin  gas  escapes  rapidly  from  its  solution  in  water,  but 
this  may  be  retarded  by  the  addition  of  a  base  transforming  it  into 
hypochlorite.  Its  oxidizing  power  is  impaired,  however,  if  the 
reaction  is  very  alkahne,  but  may  be  restored  by  bubbling  CO2 
through  the  solution." 

McClendon  emphasizes  the  importance  of  having  the  irrigating 
fluid  physiologically  normal.  It  is  not  enough  in  his  opinion  that 
the  solution  should  contain  the  salts  in  the  proper  proportion  but  it 
must  have  the  correct  hydrogen  ion  concentration  (Ph)-  This  may 
be  provided  by  bubbhng  CO2  through  the  fluid  and  measuring  the  p^ 
with  indicators. 

The  most  important  new  antiseptics  are  chloramine  T  or  sodium 
toluene  sulphonchloramide  soluble  in  water,  and  dichloramine  T  or 
toluene-p-sulphone  dichloramine  soluble  in  organic  solvents,  and  a 
paraffin  saturated  with  chlorin,  called  chlorocozane.  See  Handbook 
of  Antiseptics  (Dakin  and  Dunham).    Tr.] 

The  Method  of  Testing  Disinfectants   Considered  in  the  Light 
of  Colloid  Research. 

For  testing  disinfectants  bacteria  are  usually  dried  on  silk  threads 
or  marbles.  These  are  dipped  in  the  disinfectant  solution,  and  after 
the  solution  is  removed  they  are  placed  in  bouillon  or  fluidified  agar. 
If  the  bacteria  have  been  killed  by  the  immersion,  no  germs  develop. 


406  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

From  the  length  of  time  required  to  kill  the  germs  and  from  the  con- 
centration of  the  disinfectant  solution  we  may  judge  the  strength  of 
the  disinfectant  action. 

From  the  standpoint  of  the  colloid  chemist  the  silk  thread  pro- 
cedure contains  a  serious  error  of  method.  Even  at  present  on 
account  of  its  apparent  simplicity  this  method  is  frequently  em- 
ployed. We  know  from  practical  experience  that  silk  is  a  very 
powerful  adsorbent.  The  investigations  of  W.  Schellens  *  on  the 
relation  of  silk  to  sublimate  is  of  interest  in  this  connection.  He 
shook  1  gm.  silk  with  50  c.c.  of  1  per  cent  sublimate  solution  and 
then  determined  how  much  mercury  was  present  both  in  the  re- 
maining fluid,  and  in  the  silk  after  it  had  been  washed  many  times. 
He  found  that  the  silk  had  taken  up  6.04  per  cent  of  its  weight  of 
metallic  mercury  but  had  fixed  only  1.9  per  cent.  We  thus  see  that 
silk  retains  very  considerable  quantities  of  sublimate.  Similar  re- 
sults were  obtained  by  W.  Schellens  for  ferric  chlorid,  ferric  acetate, 
several  mercuric  salts,  lead  nitrate,  etc.  From  this  we  must  con- 
clude that  silk  is  not  a  suitable  germ  carrier  for  disinfection  experi- 
ments, since  as  the  result  of  adsorption  (no  action  can  be  ascribed 
to  the  "fixed"  mercury,  etc.)  it  retains  too  much  disinfectant;  on 
this  account  the  germ  cannot  escape  from  the  disinfectant,  and  ac- 
cordingly we  are  only  given  information  relative  to  inhibition  of 
development  and  not  concerning  the  lethal  action.  Paul  and 
Krönig  chose,  as  germ  carriers,  marbles  because  the  disinfectant 
can  barely  adhere  to  them  by  adsorption.  H.  Bechhold  and  P. 
Ehrlich,*  as  well  as  H.  Bechhold, *9  in  their  experiments  on  lethal 
action  completely  discarded  germ  carriers;  they  prepared  bac- 
terial cultures  on  agar,  which  they  covered  with  the  disinfectant 
fluid.  After  removing  the  disinfectant,  they  washed  the  culture 
twice  with  physiological  salt  solution  (which  is  finally  made  very 
faintly  alkaline)  and  then  transplanted  the  culture  to  a  new  medium 
(agar).  On  account  of  the  thickness  of  the  culture,  very  great 
demands  are  made  upon  the  disinfectant  by  this  method,  but  no 
germ  carrier  whatever  is  transferred  to  the  new  culture  medium, 
and  the  method  thus  completely  avoids  the  source  of  error  men- 
tioned above. 

The  experiments  on  the  disinfectant  action  of  formaldehyd  gave 
such  contradictory  results,  because  the  great  adsorption  of  formalde- 
hyd by  silk  was  ignored,  as  was  pointed  out,  especially  by  Schum- 

BERG.* 

In  order  to  annul  the  adsorptive  action  of  germs  and  germ  carriers 
in  disinfection  experiments,  an  attempt  was  made  to  render  the 
disinfectant  inactive  by  chemical  means,  as  it  was  found  impossible 


TOXICOLOGY  AND  PHARMACOLOGY  407 

to  accomplish  this  by  washmg.  J.  Geppert  *  inactivated  sublimate 
by  means  of  the  action  of  ammonium  sulphid ;  the  sublimate  is  thus 
changed  to  the  innocuous  mercuric  sulphid.  In  the  case  of  formal- 
dehyd,  ammonia  is  employed,  for  by  means  of  ammonia,  formalde- 
hyd  is  changed  to  hexamethylentetramin.  There  is  no  chemical 
agent  destructive  for  phenol  and  phenol-like  compounds  to  which 
objections  cannot  be  raised. 

From  the  colloid-chemical  standpoint,  I  regard  the  principle  of 
chemically  removing  the  disinfectant  as  erroneous  in  many  cases. 
The  idea  which  guided  Geppert  and  his  successors  was  evidently 
that  if  a  germ  which  had  been  immersed  in  a  disinfectant  is  placed  on 
a  suitable  culture  medium,  the  medium  abstracts  the  last  traces  of 
the  adherent  disinfectant;  it  is  thus  washed  just  as  a  chemist  washes 
a  crystalline  precipitate  on  a  filter.  In  this  way  we  consider  the 
effect  only  for  the  time  during  which  the  germ  remained  in  the  disin- 
fectant, and  J.  Geppert  and  his  followers  seek  to  imitate  this  limited 
time  by  chemical  destruction  of  the  disinfectant  when  the  germ 
is  removed.  As  a  matter  of  fact  the  process  proceeds  differently: 
when  the  germ  is  removed  from  the  disinfectant  and  is  placed  on  a 
fresh  culture  medium,  it  releases  the  disinfectant  only  slowly  and 
incompletely  in  accordance  with  the  laws  of  adsorption.  We  may 
compare  the  process  to  the  ''bleeding"  of  dyed  fabric;  especially 
the  bleeding  of  cotton  which  has  been  dyed  with  a  dye  that  is  chemi- 
cally insufl&ciently  fixed  by  the  fiber  and  which  for  days  gives  up 
color  when  washed  with  water;  the  dyer  says  it  ''  bleeds."  Thus 
for  a  long  time  by  a  pure  adsorptive  action  the  germ  retains  the 
disinfectant  and  is  injured  by  it.  That  this  assumption  is  correct  is 
shown  by  some  experimental  results  taken  from  the  literature, 
throughout  which  the  expression  is  employed  that  the  germs  are 
"weakened."  This  expression  appears  to  me  to  be  the  transfer  to 
organisms  to  which  it  no  longer  applies  of  a  conception  applicable  to 
men  and  higher  animals. 

According  to  J.  Geppert,  anthrax  spores  are  weakened  but  not 
killed  by  the  action  of  0.1  per  cent  sublimate  solution  for  15  min- 
utes. They  are  unable  to  develop  even  in  a  culture  medium  which 
contains  as  little  as  1 : 2,000,000  sublimate,  whereas  normal  an- 
thrax bacilli  thrive  quite  well  in  this  medium.  Our  interpretation 
of  this  is  that  anthrax  spores  previously  treated  with  1 :  1000 
sublimate  adsorbs  so  much  sublimate  that  they  are  in  adsorption 
equilibrium  with  a  nutrient  medium  that  contains  1:2,000,000 
sublimate. 

Heinz  says,  "Sublimate  acts  in  animal  infections  just  the  same  as 
when  transplanted  upon  artificial  media  and  the  minutest  traces 


408  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

suffice  to  prevent  multiplication  or  the  infection  of  animals  on  the 
part  of  the  germs  weakened  by  the  antiseptic. " 

''Anthrax  bacilli  (Heinz)  hke  anthrax  spores  prior  to  the  lethal 
action  show  a  stage  of  weakness  in  which  the  bacilli  are  unable  to 
grow  in  a  nutritive  medium  containing  a  minimal  amount  of  disin- 
fectant. Thus  anthrax  bacilli  which  had  been  immersed  in  1  per 
cent  carbolic  acid  (and  had  not  been  killed)  did  not  grow  in  a  culture 
medium  which  contained  a  small  amount  of  carbolic  acid, "  whereas 
fresh  anthrax  bacilli  grew  luxuriantly. 

I  find  a  very  instructive  example  in  Ottolenghi's  *  paper.  He 
says,  ''The  fact  is  very  interesting,  that  occasionally  certain  paper 
strips  (he  soaked  blotting  paper  strips  with  an  emulsion  of  anthrax 
spores,  dried  them  and  then  placed  them  in  sublimate  solution)  after 
they  have  been  subjected  for  24  hours  to  a  sublimate  solution  (up  to 
2.712  per  cent)  and  were  inoculated  into  guinea  pigs,  may  yield  a 
luxuriant  development  of  anthrax  bacilli  if  they  are  removed  from  the 
thoroughly  healthy  animal  after  one  week  and  are  placed  on  media 
after  a  thorough  treatment  with  H2S. "  The  results  of  H.  Reichen- 
bach ^  are  to  be  judged  from  the  same  standpoint.  After  treating 
anthrax  spores  with  sublimate,  they  first  lost  their  activity  in  the 
bodies  of  animals,  then  their  ability  to  grow  in  bouillon  (without 
ammonium  sulphid  treatment)  and  only  after  a  much  longer  time 
did  they  cease  to  grow,  even  after  treatment  with  ammonium  sulphid. 

Unquestionably  numerous  analogous  examples  would  be  found 
were  the  literature  carefully  studied. 

It  may  be  seen  from  this  that  in  disinfection  experiments,  the 
chemical  removal  of  the  disinfectant  may  lead  to  false  results,  that  it 
may  simulate  a  weaker  action  of  the  disinfectant  than  it  actually  has, 
i.e.,  a  weaker  action  than  it  possesses  in  practice  under  natural 
conditions.  On  this  account  I  regard  repeated  washing  of  the  germs 
with  indifferent  solvents  (water  or  physiological  salt  solution  which 
is  finally  made  faintly  alkaHne  with  soda)  as  the  proper  method  for  the 
removal  of  the  disinfectant.  Whatever  is  retained  by  the  germ  after 
such  a  washing  would  also  be  retained  under  natural  conditions.  The 
Bechhold-Ehrlich  method  of  killing  germs  (see  p.  406)  meets  all 
these  conditions  correctly. 

This  criticism  relates  to  the  testing  of  disinfectants  against  germs 
which  can  directly  enter  the  organism  (disinfection  of  the  hands, 
antiseptics,  etc.).  It  is  otherwise  with  substances  which  serve 
for  the  disinfection  of  stools,  sputum,  etc.  Under  these  circumstances 
we   must   consider   that    the   disinfectants   penetrate   an   environ- 

1  According  to  personal  letter.  See  also  H.  Reichenbach,  Zeitschr.  f.  Hy- 
giene und.  Infectionskrankh.,  50,  455,  u.  460-462  (1905). 


TOXICOLOGY  AND  PHARMACOLOGY 


409 


ment  which  contains  hydrogen  sulphid,  ammonia,  etc.  The  testing 
of  a  disinfectant  must  always  take  its  use  into  consideration  and  be 
accordingly  varied  in  different  cases.  [The  criterion  of  Carrel  and 
Dehelly  is  the  bacterial  count  per  field  in  smears  taken  from  the 
wound.     Tr.] 

Diuretics  and  Purgatives. 

Diuresis  and  defecation  may  be  influenced  in  the  most  varied  ways, 
for  instance  by  increased  blood  pressure  or  by  increased  peristalsis  — 
in  brief,  by  such  factors  as  chiefly  exert  a  more  or  less  specific  nervous 
action;  similar  effects  may  be  obtained  by  a  purely  mechanical 
facilitation  of  secretion  or  by  hindrance  of  absorption. 

We  have  repeatedly  referred  to  the  lyotropic  series  of  the  alkaline 
salts  (see  pp.  80  and  296)  and  have  shown  among  other  things,  that 
there  exists  a  remarkable  parallelism  between  the  swelling  of  gelatin 
and  fibrin,  the  precipitation  of  albumin  and  lecithin  and  the  irrita- 
bility of  frog's  muscle  and  ciliated  epithefium.  Also  for  diuresis 
and  defecation  there  exist  such  evident  relationships  which  we  shall 
here  elucidate.  We  give  the  classification  of  F.  Hofmeister.  The 
figures  above  the  columns  I,  II,  etc.,  indicate  the  concentration  of  the 
salt  solutions  which  are  necessary  to  salt  out  globulin. 


I. 

II. 

III. 

IV. 

V. 

1.51-1.66 

2-2.03 

2.51-2.72 

3.53-3.63 

5.42-5.52 

Li  sulphate 
Na  sulphate 
Na  phosphate 
K  phosphate 
K  acetate 

NH4  sulphate 

Mg  sulphate 
NH4  phosphate 
NH4  citrate 
NH4  tartrate 
Na  carbonate 

NaCl 
KCl 

Na  nitrate 
Na  chlorate 

Na  acetate 

K  citrate 

Na  citrate 

K  tartrate 

Na  tartrate 

The  various  members  of  Group  I  are  purgatives;  those  of  IV  and 
V  are  diuretics,  while  the  action  of  those  in  II  and  III  with  the 
exception  of  magnesium  sulphate  are  not  sufficiently  definite  to  be 
of  any  service. 

Obviously,  the  anion  is  of  the  greatest  importance  for  the  action 
of  the  above  salts:  we  observe  that  CI  and  NO3  have  the  highest 
rate  of  diffusion  and  are  most  rapidly  absorbed.  NaCl,  KCl  and 
NaNOs  aid  sweHing  so  that  a  gel  swells  more  rapidly  in  such  a  salt 
solution  than  in  pure  water.      From  this  it  follows  that  the  in- 


410  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

testine  will  take  up  such  solutions  more  rapidly  than  pure  water. 
Accordingly,  all  the  conditions  necessary  to  give  the  body  a  large 
quantity  of  dilute  salt  solution  are  fulfilled.  We  know  from  Chap- 
ter XIV  that  there  is  a  strong  effort  on  the  part  of  the  manmaalian 
organism  to  keep  constant  the  swollen  condition  of  the  blood  and 
tissues,  as  well  as  the  osmotic  pressure.  For  this  purpose,  the 
kidney  is  most  important,  since  it  is  able  to  remove  excess  of  water 
and  salts.  We  may  even  at  present  recognize  thus  the  quahtative 
relationship  between  physical  properties  and  the  diuretic  action  of 
Groups  IV  and  V.  Unfortunately,  we  are  not  in  a  position  to  pursue 
the  process  quantitatively,  but  we  may  assume  that  there  would  not 
be  a  simple  relationship.  The  above-mentioned  physical  properties 
of  Groups  IV  and  V  are  to  be  classified  not  only  in  reference  to  the 
intestinal  membranes  and  the  kidney  function,  but  they  also  pay 
a  role  in  the  irritation  of  nerves  and  the  contraction  of  muscle  (see 
p.  289  et  seq.  and  p.  354).  According  to  Wo.  Pauli  *^  the  majority 
of  cations  raise  the  blood  pressure,  whereas  Br  depresses  it.  This 
explains  why  bromids  are  of  no  use  as  diuretics  in  spite  of  the  fact 
that  they  might  be  classified  as  such  from  their  behavior  with  colloids; 
the  depression  of  blood  pressure  they  cause  opposes  their  diuretic 
action.  Hypotonic  common  salt  solution  and  potassium  nitrate 
solutions  remain  therefore  the  chief  diuretics  among  the  alkali  salts. 
In  fact,  it  is  the  solution  of  common  salt  which  plays,  in  the  Spa 
"mineral  water  cures,"  the  chief  part  in  increasing  the  urinary 
excretion. 

The  result  is  quite  different  when  solutions  are  introduced  directly 
into  the  blood  stream.  A  physiological  salt  solution  is  excreted 
practically  quantitatively.  If  we  inject  a  hypertonic  salt  solution, 
then  more  water  will  be  excreted  than  was  introduced,  and  (within 
certain  limits)  proportionately  more  will  be  excreted  the  greater  the 
concentration.  This  is  not  surprising,  because  the  salt  withdraws 
the  water  of  swelling,  especially  from  the  blood  corpuscles  and  the 
muscles.  The  water  thus  set  "free"  is  then  filtered  away  by  the 
kidneys.  Sulphates,  phosphates,  tartrates  and  citrates,  etc.,  of 
sodium  impede  diuresis  when  taken  by  mouth;  however,  when  di- 
rectly injected  into  the  blood  stream,  they  are  even  more  strongly 
diuretic  than  common  salt.  This  depends  on  their  strong  dehydrat- 
ing action  and  their  low  diffusibihty.  Martin  H.  Fischer  by  in- 
troducing such  salts  was  able  to  make  a  kidney,  which  had  been 
edematous  by  ligating  the  renal  artery,  function  again.  On  in- 
jecting an  appropriate  salt  into  the  renal  artery  or  even  into  the 
kidney  itself  the  swelling  subsided  and  the  anuria  ceased. 

According  to  E.  Frey,*  if  we  inject  the  salts  mentioned  along 


TOXICOLOGY  AND  PHARMACOLOGY  411 

with  narcotics  (morphine,  chloral,  ether,  urethan),  no  diuresis  de- 
velops, and  on  the  other  hand  the  absorption  of  water  from  the  in- 
testines is  unimpaired.  This  is  explained  by  the  fact  mentioned 
(p.  338),  that  such  narcotics  inhibit  the  oxidizing  processes  in  the 
organism,  which  results  in  a  greater  fixation  of  water  ("acid  swell- 
ing"). 

What  holds  for  electrolytes  is  also  true  for  nonelectrolytes.  We 
have  recognized  in  urea  a  substance  which  greatly  aids  diffusion 
through  jellies  (see  p.  55)  and  which  opens  through  the  hydrogel 
paths  for  itself  and  other  substances;  as  a  matter  of  fact  it  acts  as 
a  diuretic.  I  wish  to  mention  some  additional  facts  concerning 
ammonium  salts  and  the  cleavage  products  of  protein.  All  the  evi- 
dence (see  pp.  80  to  82)  is  in  favor  of  the  view  that  the  action  of 
the  cations  and  of  the  anions  of  an  electrolyte  is  antagonistic  and  that 
they  mutually  counteract  a  portion  of  their  own  activity.  Thus  NH4 
seems  to  oppose  the  precipitating  and  dehydrating  action  of  SO4, 
citrate,  and  tartrate  anions  to  a  greater  extent  than  K  and  Na  (see 
the  Series  III  of  our  group).     If  we  bring  this  into  relation  with 

/NH2 

analogous  action  of  urea  CO  <f  we  may  m  general  attnbute  to 

XNHa 

the  NH2  and  NII3  groups  the  property  of  aiding  diffusion  and  we 

also  understand  the  ease  with  which  protein  cletivage  products  are 

absorbed,  for  they  occur  in  the  intestines  largely  spht  into  substances 

with  free  NH  and  NH2  groups. 

Maktin  H.  Fischer  explains  the  diuretic  action  of  digitalis  prepa- 
rations and  of  caffein  as  follows.  They  increase  the  strength  and 
frequency  of  the  pulse,  increase  the  utiHzation  of  oxygen  and  thus 
the  blood  supply  of  the  kidneys  is  increased  and  the  ''free"  water  in 
the  blood  is  increased  and  may  be  excreted.  This  agrees  with  the 
results  of  Sobieranski,  Hirokowa  and  Grünwald  who  found  that 
the  diuretic  action  of  caffein,  theohromin  and  diuretin  depended  on 
their  interference  with  reabsorption. 

Grünwald  was  able  to  show,  for  instance,  that  rabbits  on  a 
chlorin-free  diet  when  treated  with  theobromin  finally  perished  for 
want  of  chlorin.  The  chlorin  removed  from  the  body  by  ultrafiltra- 
tion was  not  restored  by  reabsorption. 

Purgatives. 

If  we  wish  to  explain  the  action  of  purgatives  we  must  first  re- 
view the  processes  in  the  intestines.  The  intestine  is  the  place  where 
secretion  and  absorption  occur.  The  volume  of  the  secretion  of  the 
salivary  glands,  the  stomach,  bile,  pancreas  and  intestine  is,  accord- 


412  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

ing  to  H.  Meyer  and  R.  Gottlieb,*  about  3  to  4.5  liters  daily.  The 
amount  reabsorbed  is  a  still  larger  quantity.  An  increased  absorp- 
tion of  fluid  is  followed  by  an  increased  secretion  of  fluid  into  the 
intestine.  The  final  result  depends  on  whether  more  fluid  is  ab- 
sorbed or  secreted  (into  the  intestine)  or  vice  versa.  If  the  secretion 
exceeds  reabsorption  the  intestinal  contents  are  fluid  and  volumi- 
nous, and  we  have  one  of  the  conditions  for  easy  defecation.  On  this 
account  substances  having  a  capacity  for  swelling  counteract  con- 
stipation. Persons  suffering  from  constipation  are  recommended  to 
eat  considerable  quantities  of  vegetables  and  graham  bread,  because 
the  indigestible  cellulose  they  contain  retains  water.  This  accounts 
for  the  laxative  action  of  agar.  In  addition,  all  substances  which  act 
on  the  intestinal  nerves  by  increasing  peristalsis  will  favor  defecation. 

The  effect  of  alkali  salts  have  been  most  exhaustively  investigated, 
but  before  we  consider  them  we  must  recall  the  observations  of 
LoEPER.*  He  found  that  salt  solutions  which  were  introduced  orally 
either  in  hypertonic  or  in  hypotonic  solution,  when  they  reached  the 
intestine  were  in  practically  isotonic  solution.  We  can  accordingly 
disregard  all  hypotheses  which  seek  to  explain  the  action  of  purgatives 
by  differences  of  the  osmotic  pressure  of  the  intestinal  contents.  If 
hypertonic  or  hypotonic  salt  solutions  have  an  effect  notwithstanding, 
we  must  explain  this  by  indirect  action,  for  hypertonic  salt  solutions 
inhibit  gastric  movements  and  thus  interfere  with  the  progress  of 
the  chyme  from  stomach  to  intestine. 

We  saw  that  the  chlorids  and  nitrates  are  diuretics;  the  sulphates, 
phosphates,  citrates  and  tartrates  are  chiefly  purgatives,  so  that  the 
last-mentioned  anions  must  possess  properties  which  either  increase 
secretion,  diminish  absorption,  or  strengthen  peristalsis. 

Some  diuretics  may  purge  by  reason  of  increased  secretion.  Table 
salt  acts  in  this  way  and,  in  mild  constipation,  it  is  given  in  dry 
form  in  Spa  cures,  or  as  sodium  bicarbonate.^ 

In  the  case  of  the  real  purgatives  of  Group  I,  it  is  a  question 
whether  their  action  is  directly  on  the  intestinal  mucous  membrane 
or  whether  they  increase  peristalsis  through  nerve  stiniulation.  Pos- 
sibly they  may  impede  the  absorption  of  themselves  and  other  sub- 
stances by  dehydrating  and  precipitating  albumin.  This  does  occur 
in  high  concentration  (1  gram  equivalent  Na2S04).  G.  Quagliari- 
ELLO  *  has  shown  in  the  case  of  sodium  sulphate  that  for  such  salt 
concentrations  as  enter  the  intestine  after  passing  through  the 
stomach,  the  imbibition  of  water  is  no  different  than  for  sodium 
chlorid,  and  accordingly  that  there  is  no  direct  action  by  such  purga- 
tive salts  on  the  intestinal  mucous  membrane. 

^  Changes  into  sodium  chlorid  with  the  gastric  hydrochloric  acid. 


TOXICOLOGY  AND  PHARMACOLOGY  413 

We  must  therefore  strive  to  discover  Aviiat  facts  speak  for  a 
strengthening  of  peristalsis  by  such  salts. 

Interesting  observations  by  MacCallum*  in  agreement  with 
observations  by  J.  Loeb  *'  show,  as  a  matter  of  fact,  that  the  salts 
of  Group  I  exert  on  muscle  and  nerve  a  stimulating  effect  which 
induces  an  increased  peristalsis  of  the  intestines.  This  occurs  not 
only  when  citrates,  tartrates  and  sulphates  are  placed  in  the  lumen 
of  the  intestine  but  also  when  they  are  injected  subcutaneously  or 

intravenously,  and  even  dropping  a  —  solution  of  such  salts  on  the 

o 

peritoneal  surface  of  the  intestines  induces  especially  strong  peris- 
taltic intestinal  movement,  so  that  according  to  Wo.  Pauli*''  the 
intestinal  activity  may  even  approach  that  of  a  gastro-enteritis. 
With  this  increase  of  peristalsis  there  is  associated  an  active  secre- 
tion into  the  intestine,  so  that  according  to  MacCallum  the  empty 
coils  of  small  intestine  of  a  rabbit  became  filled  with  secretion 
(20  c.c.)  when  a  drop  of  sodium  citrate  solution  was  placed  on  the 
peritoneal  coat.  I  wish  to  recall  that  those  anions  raise  the  blood 
pressure,  and  possibly  the  increased  secretory  activity  stands  in 
relation  to  this. 

Magnesium  sulphate  is  one  of  the  best  known  cathartics,  although 
on  account  of  its  place  in  Group  III  of  our  table  we  would  hardly 
expect  that  it  should  have  any  special  action.  This  need  not  surprise 
us,  since  there  are  in  the  intestine  Na  ions  which  largely  inhibit  the 
antagonistic  action  of  Mg  ions,  and  as  a  result  the  SO4  action  is 
brought  out.  According  to  MacCallum,  if  we  introduce  MgCl2 
(instead  of  MgS04)  or  CaCl2  solution  into  the  intestine,  or  inject 
them  into  the  circulation,  the  peristal  ic  waves  which  citrates  or 
fluorids,  for  instance,  strongly  induce,  are  inhibited.  This  agrees 
completely  with  our  premises,  according  to  which  there  exists  high 
antagonistic  action  of  divalent  cations  (see  p.  82)  as  here  exemplified. 
CaCl2  diminishes  diuresis  as  well  as  defecation. 

Frankl  *  and  Auer  *  found  certain  contradictions  to  the  results 
of  MacCallum.  They  claim  to  have  observed  no  diarrhea  upon 
injecting  subcutaneously  or  intravenously  dilute  purgative  salts,  and 
to  have  observed  even  constipation  upon  employing  more  concen- 
trated solutions.  J.  Bancroft,*  on  the  contrary,  confirms  the  results 
of  MacCallum.  From  all  of  which  it  may  be  concluded  that,  as 
was  to  be  expected,  the  result  chiefly  depends  upon  the  conditions 
of  concentration  and  upon  the  location  where  these  conditions  are 
active.  In  this  connection  the  old  experiments  of  von  Hay*  are 
very  instructive.  If  he  gave  large  quantities  of  sodium  sulphate  by 
mouth,  it  abstracted  fluid  until  its  concentration  had  fallen  to  3  per 


414  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

cent  after  which  diarrhea  occurred.  If  he  deprived  the  animal  of 
water  for  1  or  2  days  and  gave  only  a  dry  diet,  even  concentrated 
solutions  of  Glauber's  salt  (20  gm.  salt)  had  no  cathartic  action. 
The  same  quantity  of  salt  diluted  to  a  5  per  cent  solution  resulted  in 
strong  catharsis  after  one  or  two  hours.  We  see,  therefore,  that  the 
condition  of  the  tissues  and  blood  colloids  in  respect  to  swelling  play 
a  most  important  part  in  these  processes.  On  this  account  it  is 
possible  to  employ  the  salts  of  Group  I  and  MgS04  either  to  purge  or 
to  dehydrate  the  body  as  may  be  necessary.  It  is  important  to  con- 
sider how  purgative  action  is  to  be  measured;  whether  according  to 
the  amount  of  dried  substance  passed  or  according  to  the  total 
quantity  including  the  fluid.  A  fluid  stool  permits  us  to  conclude  that 
there  is  an  increase  of  secretion  or  a  diminution  of  absorption,  an 
increased  amount  of  solid  feces  suggests  an  increase  of  peristalsis. 


Astringents. 

Constipating  substances,  i.e.,  those  which  diminish  the  intestinal 
or  even  the  gastric  secretion  and  peristalsis,  are  substances  which 
diminish  stimuli.  As  such  are  employed,  as  has  been  mentioned  on 
page  365,  hydrophile  colloids  (mucilages,  etc.),  as  well  as  strongly 
adsorbent  suspensions  (talc,  bismuth  subnitrate,  bismuth  subgallate). 
More  powerful  actions  are  obtained  with  such  substances  which  tan 
the  intestinal  membrane  superficially  and  in  this  way  interfere  with 
the  secretion  of  the  intestinal  glands,  or  at  least  arrest  absorption 
at  the  point  affected.  Most  important  of  these  is  tannin  and  such 
tannin  compounds  as  are  dissolved  in  the  intestinal  juice  (tannalbin, 
tannigen). 

BALNEOLOGY. 

It  would  certainly  be  a  fortunate  circumstance  for  balneology  if 
we  knew  but  a  small  fraction  as  much  about  the  physiological  action 
of  medicinal  springs  as  we  at  present  know  of  their  physical  and 
chemical  properties.  Every  mineral  spring,  no  matter  how  insignifi- 
cant, has  its  chemical  composition  determined  to  the  fifth  decimal ;  its 
osmotic  pressure,  conductivity,  radioactivity,  etc.,  are  investigated 
and  the  possibilities  of  its  therapeutic  activity  are  lauded  and  its 
clinical  successes  (not  the  failures)  are  carefully  registered. 

The  scientifically  determinable  and  explicable  effects  are  con- 
sidered by  very  few.  From  these  remarks  it  must  not  be  concluded 
that  valuable  clinical  results  are  not  to  be  obtained  by  means  of 
balneology,  but  that  its  scientific  basis  is  largely  undetermined. 


TOXICOLQGY  AND  PHARMACOLOGY  415 

Water  and  Solutions. 

In  Chapter  XIV I  stated  that  the  body  colloids  have  a  normal  state 
of  swelling  and  that  they  stand  in  definite  swelling  relations  to  each 
other.  Thus,  when  the  condition  of  swelhng  in  one  organ,  i.e.,  the 
muscles,  changes,  it  must  in  turn  influence  the  condition  of  swelling 
of  other  body  colloids;  as  in  the  case  of  every  other  substance  there 
is  for  water,  a  definite  distribution  in  the  organism.  This  distri- 
bution of  water  is  dependent  on  the  capacity  of  the  organ  colloids 
to  swell  and  this  again  largely  depends  on  the  amount  of  electrolytes 
contained. 

It  is  very  probable  that  during  life  the  crystalloid  content  of 
organ  colloids  suffer  certain  changes  which  may  even  become  patho- 
logical. Under  such  circumstances,  it  is  conceivable  that  a  thorough 
flushing  of  the  body  with  water  such  as  our  ancestors  were  accustomed 
to  undertake  every  spring  to  "purify  the  blood"  might  be  of  great 
value  inasmuch  as  it  restores  the  normal  swelling. 

It  would  be  very  desirable  for  the  elucidation  of  this  question  to 
undertake  a  thorough  experimental  investigation  of  the  swelling 
capacity  and  swelling  range  of  the  organs  at  various  ages  under 
normal  and  pathological  conditions. 

Just  as  a  drinking  "cure"  so  a  thirst  "cure"  (Schroth's  "cure") 
may  influence  the  condition  of  swelling.  [Karell's  Treatment  as 
well  as  Tuffnel's  owe  much  of  their  efficacy  to  "  drink  restric- 
tion."    (See  p.  234.)     Tr.] 

In  various  parts  of  this  book,  there  have  been  thoroughly  de- 
scribed the  great  significance  of  electrolytes  for  the  swelling  of  cell 
colloids,  the  viscosity  of  the  blood,  the  influence  of  heavy  metals  on 
solubility  (urates,  lime  salts)  and  the  acceleration  of  fermentative 
cleavages  and  syntheses,  i.e.,  the  acceleration  of  metabolism.  The 
introduction  into  the  body  of  electrolytes  in  the  form  of  mineral 
waters  is  the  original  and  essential  function  of  balneology.  It  is 
quite  conceivable,  that  the  increase  in  the  body  of  definite  anions 
or  cations  might  satisfy  important  therapeutic  demands.  [Radio- 
activity should  be  considered  a  possible  therapeutic  adjuvant,  see 
ZwAARDEMAKER,  H.,  Aequi-radioactivity,  Am.  Jour.  Physiol.,  1918, 
XLV,  p.  147.    Tr.] 

We  have  at  present  nothing  to  support  even  the  idea,  that  concen- 
tration of  a  definite  electrolyte  is  possible. 

There  are  a  few  indications  which  favor  this  view.  It  may  be  re- 
called, that  hypertonic  salt  solutions  result  in  a  destruction  of  tissue; 
hypotonic  solutions,  in  a  diminished  metabolism  of  protein  (see  E. 
RosT  *) .     We  must  also  remember  that  cells  are  not  impermeable  for 


416  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

ions,  but  that  accompanying  the  abstraction  of  water  by  hypertonic 
solutions  a  penetration  or  interchange  of  ions  may  occur. 

For  the  diuretic  and  purgative  action  of  neutral  salts  see  page 
409  et  seq. 

SALVES,  LINIMENTS. 

Salves  and  similar  preparations  are  frequently  employed  not  only 
to  cover  wounds  but  also  with  the  object  of  introducing  medicines 
into  the  body  through  the  skin.  The  skin  absorbs  only  such  sub- 
stances as  are  soluble  in  fat;  this  has  been  established  by  the  inves- 
tigations of  W.  FiLEHNE*  and  A.  Schwenkenbecher.* 

The  colloidal  properties  of  fats  deserve  our  attention.  The  cooling 
sensation  induced  by  cold  cream  depends  on  its  capacity  for  holding 
water;  it  takes  up  about  28  per  cent  water  which  is  obviously  the 
dispersed  phase.  Wool  fat  is  "hydrophile"  to  a  still  greater  extent. 
It  enters  commerce  as  lanolin  and  is  the  base  for  salves.  We  saw 
that  by  means  of  hydrophile  lecithin,  water-soluble,  though  ordinarily 
fat-insoluble,  substances,  such  as  sugar,  become  soluble  in  fat.  Pos- 
sibly we  may  attribute  to  this  property,  when  it  is  present,  the 
penetration  into  the  body  through  the  skin  by  means  of  salves,  of 
medicaments  for  which  the  skin  is  otherwise  impermeable.  [Ax- 
ELROD,  Jour.  Industr.  and  Eng.  Chem.,  Vol.  IX.,  p.  1123,  gives  the 
method  for  preparing  cetyl  alcohol  as  a  substitute  for  lanolin  and 
eucerin.     Tr.] 

P.  G.  Unna*  showed  that  the  hydrophile  constituent  of  wool  fat 
is  the  oxy Cholesterin  group.  Five  parts  of  this  latter,  mixed  with 
95  parts  of  paraffin  ointment,  are  able  to  unite  with  100  per  cent  of 
water.     (It  enters  commerce  as  eucerin.) 


X 


CHAPTER  XXIII. 

MICROSCOPICAL   TECHNIC. 

The  microscopic  study  of  organisms  and  parts  of  organs  is  one  of 
the  most  important  tasks  of  biologists  and  physicians.  The  princi- 
pal object  of  the  microscopist  is  to  deduce  from  the  form  of  an  object 
its  nature  and  whether  its  appearance  is  normal  or  diseased. 

Remarkable  progress  has  been  made  in  this  field  though  the  chemi- 
cal interpretation  of  the  methods  employed  is  still  in  its  infancy. 

To  prepare  an  object  for  microscopical  examination  it  must  be 
spread  out  very  thin  upon  a  slide  and,  if  necessary,  made  transpar- 
ent. In  the  case  of  unicellular  organisms  (bacteria,  protozoa,  etc.) 
no  further  preparation  is  necessary.  If  the  presence  of  bacteria  is 
to  be  determined  it  suffices  to  spread  the  object  in  question  on  a 
slide  ^^dth  a  platinum  loop  and  to  dry  it  at  moderate  temperature 
by  drawing  the  slide  through  a  Bunsen  flame  several  times,  so  as  to 
coagulate  the  albumin.  On  account  of  the  intensity  with  which 
bacteria  and  cocci  stain  with  basic  dyes  (methylene  blue,  carbol 
fuschin,  etc.)  it  is  usually  an  easy  matter  to  recognize  them  in  the 
otherwise  structureless  coagulum. 

Organs  of  higher  plants  and  animals  must  be  either  teased  or  pre- 
pared in  thin  sections. 

The  least  deceptive  object  is  naturally  the  living  organism,  as  we 
see  it,  for  instance,  in  hanging  drops,  in  the  moist  chamber  or  the 
microscope  stage  aquarium  of  J.  Cori,  etc.;  even  in  higher  ani- 
mals some  investigations  may  be  made  while  they  are  still  aUve  by 
spreading  out  portions  of  organs  still  connected  with  the  animal  so  that 
they  are  transparent.  In  this  way  we  may  see,  for  instance,  the  cir- 
culation of  the  blood  in  the  lungs  and  in  the  web  of  a  frog's  foot.  Much 
more  frequently  an  opportunity  to  examine  sur\aving  tissue  pre- 
sents itself.  It  is  by  no  means  necessary  that  at  the  moment  that 
the  animal  itself  dies,  a  given  organ  or  cell  should  die.  Let  me  recall 
that  the  heart  may  be  isolated  immediately  after  an  animal  (cat, 
frog,  etc.)  has  been  killed  and  may  continue  to  beat  for  a  long  time 
if  suitable  means  are  employed.  Leucocytes  of  warm  blooded 
animals  show  protoplasmic  movements,  if  observed  at  37°,  even  as 
long  as  a  half  day  after  the  animal's  death.  It  goes  without  saying 
that  this  must  occur  in  a  medium  which  causes  neither  swelling  nor 

417 


418  '        COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

shrinking.  Pure  water  is  never  suited  for  this;  a  salt  solution 
which  approaches  in  its  composition  that  which  bathes  the  organ  is 
best.  In  many  cases  "physiological  salt  solution"  is  ample;  in 
mammals  this  contains  0.85  per  cent  common  salt,  in  other  kinds  of 
animals  this  may  advantageously  be  reduced  to  0.5  per  cent.  Even 
though  the  living  or  surviving  organism  presents  a  microscopical 
picture  devoid  of  artifacts,  on  the  one  hand  it  is  rarely  possible  to 
examine  it  and,  on  the  other,  many  details  are  concealed,  since  the 
refraction  of  the  different  cell  elements  is  almost  equal.  On  this 
account  we  are  compelled  to  section  and  stain  the  organs. 

Since  even  the  death  of  the  cell  causes  changes  in  structure,  these 
changes  with  the  chemical  manipulations  about  to  be  described 
may  reach  a  grade  which  leads  to  the  gravest  errors.  Only  absolute 
ignorance  of  colloid  processes  explains  how  artificially  produced 
flocculations,  coagulations  and  striations  (after  treatment  with 
silver  nitrate  and  potassium  bichromate),  etc.,  could  be  considered 
definite  constituents  of  cells,  and  it  is  a  pity  that  numerous  pains- 
taking investigations  must,  as  a  result  of  this,  be  considered  mere 
waste  paper.  By  indicating  these  errors  A.  Fischer  and  Walther 
Berg,  as  well  as  Th.  v.  Wasielewski,  performed  a  great  service. 
Accordingly,  A.  Fischer  distinguishes  reagents  which  form  granules 
(nuclei)  and  those  which  form  coagula,  and  W.  Berg  also  calls  at- 
tention to  those  which  produce  granulated  pellicles  and  cavities. 

If  then,  as  may  be  seen  from  the  foregoing,  we  may  obtain  different 
structures  by  means  of  different  reagents  acting  on  the  same  bodies 
and,  conversely,  with  the  same  chemical  substance  produce  the  same 
microscopic  picture  on  different  bodies,  we  may  by  accurate  com- 
parative experiments  make  valuable  deductions.  It  is  hardly  possible 
to  employ  such  methods  in  determining  form,  but  for  the  understand- 
ing of  the  colloidal  nature  of  the  object  examined  they  offer  a  broad, 
uncultivated  and  promising  field  for  colloid  research. 

The  preparation  of  dead  material  for  microscopic  examination  may 
be  divided  into  maceration  and  isolation,  fixation  and  hardening, 
decalcification,  bleaching,  embedding,  sectioning  and  mounting,  and 
finally  staining.^ 

Maceration  and  Isolation. 

Maceration  and  isolation  are  for  the  purpose  of  dissolving  apart 
the  constituents  of  an  organ  (cells)  and  thus  to  recognize  their  con- 
nection and  to  make  it  possible  to  examine  the  isolated  cells.     The 

^  I  have  for  the  most  part  followed  here  the  directions  given  in  the  "Lehrbuch 
der  Mikroskopischen  Technik"  of  Prof.  Dr.  B.  Grawitz  (Leipzig,  1907),  whose 
numerous  detailed  directions  should  be  of  valuable  assistance  to  every  biochemist. 


MICROSCOPICAL   TECHNIC  419 

objects  are  placed  (depending  on  the  variety)  in  15  to  35  per  cent 
alcohol  or  in  physiological  salt  solution  which  contains  2  cc.  of  40  per 
cent  formaldehyd  per  liter,  or  into  0.1  per  cent  to  0.005  per  cent 
chromic  acid,  or  1  per  cent  osmic  acid,  dilute  picric  acid,  20  per  cent 
nitric  acid,  pure  HCl,  javelle  water  or  many  other  solutions  recom- 
mended for  particular  purposes.  Obviously,  with  the  first  mentioned 
substances  we  may  dissolve  the  connections  by  a  differential  shrinking 
of  the  cell  constituents,  because  as  we  shall  see  the  identical  substances 
are  employed  in  different  concentrations  for  fixation  and  for  harden- 
ing. Substances  such  as  20  per  cent  nitric  acid,  pure  HCl,  etc., 
obviously  change  the  cement  substances  chemically.  We  understand 
the  action  of  digestive  fluids  (pepsin-HCl,  pancreatin)  to  be  similar, 
yet  they  have  not  proven  very  satisfactory.  After  successful  "mac- 
eration" it  is  sometimes  sufficient  to  shake  violently  the  object  which 
has  been  treated  to  cause  it  to  fall  apart  or  it  may  be  teased  on  the 
slide  with  needles  or  a  coarse  brush. 


Fixing  and  Hardening. 

Whereas  the  methods  previously  described  permit  the  recognition 
of  individual  elements  of  a  tissue,  they  do  not  permit  a  study  of  the 
relations  of  the  tissue  elements,  their  connections  and,  in  short,  the 
entire  tissue  structure.  For  this  purpose  a  thin  section  of  tissue 
must  be  prepared  and  stained.  Before  doing  this  it  is  frequently 
necessary  to  fix  and  harden  the  object  to  be  studied. 

Fixation  is  undertaken  for  the  purpose  of  making  the  partly  fluid 
and  partly  semifluid  constituents  firm,  so  that  they  stop  changing, 
neither  swelling,  shrinking,  coagulating  nor  the  like  and  so  that  their 
appearance  shall  remain  as  nearly  lifelike,  or  at  least  as  fresh  as 
possible,  in  order  that  this  condition  shall  be  maintained  through  all 
the  later  manipulations.  By  fixation,  the  relations  between  the  dif- 
ferent tissue  elements  are  made  permanent.  The  object  is  frequently 
too  soft  to  section,  so  that  it  must  be  subjected  to  a  special  proce- 
dure, —  hardening.  Viewed  colloid-chemically,  tissues  may  be  consid- 
ered to  consist  of  (1)  irreversible  slightly  elastic  gels,  (2)  reversible 
elastic  gels,  (3)  sols.     All  sorts  of  transition  states  exist. 

Accordingly,  fixation  renders  each  constituent  completely  insol- 
uble, unshrinkable  and  incapable  of  swelling;  the  sols  are  changed 
to  gels,  and  no  shrinking  or  swelling  should  occur  during  the  fixation. 
Finally,  it  must  be  possible  to  stain  the  objects  well,  and  'conse- 
quently their  chemical  properties  must  not  be  too  radically  changed. 
It  is  a  problem  almost  impossible  to  solve.  For  the  sake  of  compari- 
son, every  one  knows  how  great  and  almost  insuperable  difficulties 


420  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

are  often  presented  by  the  fusing  together  of  a  metal  wire  and  a  glass 
tube  because  of  the  different  degree  of  contraction  (shrinking)  upon 
cooling.  Cracks  frequently  occur.  Now  imagine  how  very  compli- 
cated the  problem  becomes  whenever  three,  four  or  perhaps  a  dozen 
fused  ingredients  are  subjected  to  a  manipulation  to  which  they  react 
differently.  This  simple  consideration  teaches  us  that  we  can  never 
expect  a  piece  of  tissue  that  is  fixed  and  hardened  to  show  the  same 
appearance  as  when  it  is  alive.  Only  by  comparing  pieces  of  tissue, 
treated  in  different  ways,  can  we  recognize  what  is  normal  and  what 
is  due  to  fixation,  but  even  in  these  distorted  pieces  we  can  see  the 
places  of  least  resistance  where  inequalities  of  staining  exist,  and  by 
careful  consideration  we  can  learn  much  even  from  such  pieces  as 
are  regarded  as  spoiled  by  histologists. 

Of  course,  a  fixative  must  not  block  its  own  path.  If  massive 
organs,  e.g.,  the  brain  or  liver,  are  to  be  fixed,  the  fixative  solutions 
have  a  great  distance  to  travel  before  they  reach  the  center;  if 
shrinkages  or  precipitates  are  formed  at  the  periphery,  the  diffusion 
paths  are  closed  at  the  outset  and  the  fixative  could  never  reach  the 
center  even  if  such  objects  should  lie  in  the  fluid  for  weeks.  It  is 
quite  reasonable  to  expect  that  in  such  large  organs  the  central 
portion  will  show  a  different  kind  of  fixation  than  the  periphery. 

We  can  well  understand  how  temperature  plays  an  important  role, 
conditioning  not  only  the  rate  of  diffusion  but  also  governing  the 
processes  of  coagulation. 

Wherever  feasible  the  objects  are  cut  into  small  pieces  and  placed 
in  very  large  quantities  of  fixative  fluids  (50-100  times  the  volume  of 
the  object)  so  that  too  great  a  dilution  shall  not  occur  and  the  action 
shall  be  quite  uniform;  if  the  fixation  is  prolonged  the  fluid  must  be 
renewed  from  time  to  time. 

Among  the  fixatives  an  important  role  is  played  by  certain  elec- 
trolytes (chromic  acid,  bichromate,  mercuric  chlorid,  picric  acid,  etc.) ; 
they  cause  swelling  in  solutions  that  are  too  dilute,  shrinking  in  too 
concentrated  solutions.  On  this  account  C.  Dekhuysen  and  W. 
Stoeltzner  prepared  "isotonic"  solutions  which  cause  neither  swell- 
ing nor  shrinking.  These  authors,  as  may  be  seen  from  their  method 
of  expression  (hypertonic,  hypotonic),  evidently  proceeded  from  prem- 
ises which  depend  upon  osmotic  pressure,  a  factor  involved  only  to  a 
very  limited  extent.  By  treating  the  whole  practice  of  fixation  from  a 
colloid  standpoint,  doubtless  a  whole  series  of  valuable  new  methods 
of  fixation  would  be  evolved  for  histologists.  Such  a  study  would 
furnish  us  with  more  definite  rules  for  knowing  why,  on  the  one 
hand,  one  solution  is  more  suitable  for  marine  animals,  and  on  the 
other,  why  other  solutions  are  more  suitable  for  mammalian  organs. 


MICROSCOPICAL   TECHNIC  421 

It  is  quite  clear  that  the  action  of  the  same  fixative  differs,  depend- 
ing upon  the  variation  of  the  electrolyte  content  of  various  animals 
and  plants. 

From  what  has  been  said,  it  may  be  concluded  that  alkaline  solu- 
tions are  not  to  be  considered  fixatives,  since  they  produce  only 
swelling.  The  salts  of  the  light  metals  are  not  included  among  fixa- 
tives; they  usually  form  reversible  gels;  but,  on  the  contrary,  certain 
heavy  metals  as  well  as  acids,  especially  the  acid  mixtures,  are  of 
great  importance.  Many  have  an  oxidizing  action,  as  a  result  of 
which  the  organic  substances  lose  their  ability  to  swell. ^ 

Acids  are  obviously  employed  with  the  object  of  changing  sols 
into  gels,  and  since  a  chemical  change  must  simultaneously  occur,  it 
follows  that  all  acids  are  not  available  for  this  purpose  and  that  they 
must  always  be  employed  in  high  concentration.  Hydrochloric  acid 
and  sulphuric  acid  (the  latter,  at  least,  never  unmixed)  are  not  em- 
ployed as  fixatives,  but  we  do  frequently  employ  nitric  acid  in  2  to 
10  per  cent  concentration.  Its  use  is  not  general  since  it  frequently 
alters  the  stains.  For  organs  with  epidermal  coverings,  nitric  acid 
is  unsuitable  since  it  raises  the  epithelium  in  blisters  from  the  tissues 
supporting  it. 

Chromic  acid  (introduced  by  Hannover  in  1840)  is  the  oldest  and 
most  used  fixative  and  hardening  agent  for  cell  protoplasm  and  nucleus. 
It  is  used  in  concentrations  of  from  0.33  per  cent  up  to  1  per  cent. 
It  is  employed  preferably  in  the  dark,  since  daylight  causes  a  sort  of 
tanning  of  the  periphery  so  that  the  chromic  acid  penetrates  very 
slowly  and  amorphous  deposits  form  very  easily  in  the  preparation. 

Objects  fixed  in  chromic  acid  or  its  salts  become  green  in  time 
(reduction  to  chromic  oxid)  and  are  poorly  stained.  Many  methods 
for  regenerating  the  staining  capacity  of  such  specimens  have  been 
proposed  (L.  Edinger  and  Mayer,  B.  Grawitz). 

Osmic  acid  (0.5  to  2  per  cent)  is  especially  recommended  for  the 
fixation  of  protoplasm  and  nucleus.  It  is  soluble  in  fat  and  conse- 
quently can  penetrate  the  living  cell.  It  does  not  penetrate  far, 
however,  and  on  this  account  is  available  only  for  small  or  thin 
objects.  With  the  fixation  there  is  a  blackening,  especially  of  the 
fats  and  some  other  substances  (reduction  to  colloidal  osmium). 
Opinions  on  the  use  of  osmic  acid  are  very  divergent.  Though 
praised  by  some,  A.  Fischer,  as  the  result  of  his  studies  of  non- 
biological  material,  considers  it  a  weak,  unsatisfactory  precipitat- 
ing agent,  since  it  precipitates  only  acid  reacting  structures. 

1  To  fix  tissue  practically,  the  exact  directions  as  they  are  found  in  the  books 
must  be  followed;  they  are  employed  just  as  a  cooking  recipe  would  be.  Only  a 
few  generalizations  can  be  given  here. 


422  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Acetic  acid  alone  (in  concentration  up  to  1  per  cent)  causes  swelling, 
but  is  suitable  for  combination  with  reagents  which  cause  shrinking, 
especially  for  the  fixation  of  nuclear  structures. 

Trichloracetic  acid  (5  per  cent  to  10  per  cent)  penetrates 
rapidly,  destroys  the  most  delicate  structural  relations  of  pro- 
toplasm and  nucleus  but  fixes  well  centrosomes,  chromosomes  and 
spindles.  Since  fibrillar  connective  tissue  swells  strongly  in  tri- 
chloracetic acid,  the  preparation  must  be  placed  at  once  in  absolute 
alcohol. 

Picric  acid  does  not  change  all  sols  and  reversible  gels  into  irre- 
versible gels.  This  follows  from  the  results  of  A.  Fischer  who 
showed  that  precipitations  with  picric  acid  are  dissolved  again  by 
water;  and  this  is  confirmed  by  the  experience  of  histologists  with 
actual  specimens.  Only  when  combined  with  other  acids  (acetic 
acid,  chromic  acid,  sulphuric  acid,  nitric  acid  and  osmic  acid)  does 
picric  acid  attain  its  importance  as  a  fixative,  and  under  these  cir- 
cumstances it  is  highly  praised. 

Since  picric  acid  is  lipoid  soluble  and  forms  insoluble  dye  salts 
with  dye  bases  it  is  suitable  for  fixing  specimens  after  ''vital  stain- 
ing." 

Salts.  Among  these,  the  chlorids  next  to  the  Chromates  enjoy 
especial  popularity.  1  attribute  this  to  their  ease  of  diffusion  and  to 
the  fact  that  in  respect  to  swelling  and  shrinking,  the  chlorin  ion 
occupies  approximately  a  middle  position. 

Copper  chlorid  and  copper  acetate  are  suitable  for  delicate  lower 
plants  but  are  very  seldom  employed. 

Mercuric  chlorid  in  concentrated  aqueous  solution  is  very  suitable 
for  the  fixation  of  animal  preparations.  I  have  had  very  good  re- 
sults in  fixing  leucocytes.  It  may  not  be  employed  for  any  molluscs 
or  for  fresh  water  crustaceans;  it  also  seems  not  quite  suitable  for 
plant  cells.  As  a  result  of  a  certain  amount  of  lipoid  solubility  it  is 
able  to  penetrate  the  living  cells  and  on  this  account  in  vital  stain- 
ing it  serves  to  fix  the  dye.  Obviously,  part  of  the  mercuric  chlorid 
is  in  this  case  bound  by  the  protoplasmic  albumin;  the  resulting 
combination  is  somewhat  insoluble  in  water. 

Ferric  chlorid  in  alcoholic  solution  is  recommended  for  pelagic 
marine  animals. 

Platinum  chlorid  (0.1  to  1  per  cent),  palladious  chlorid  (0.1  per 
cent)  and  iridium  chlorid  are  recommended  for  special  purposes  (usu- 
ally in  mixtures). 

Potassium  bichromate  is  rarely  used  as  a  pure  solution,  since  it 
markedly  changes  the  structure,  but  used  in  combination  with  other 
substances  it  is  a  very  popular  fixative  (with  acetic  acid  for  cell  sub- 


MICROSCOPICAL   TECHNIC  423 

stance  and  nuclear  structure;  with  sodium  sulphate  for  the  central 
nervous  system;  with  copper  sulphate  for  bulky  objects;  with 
sublimate,  etc.). 

Nonelectrolytes. 

Alcohol.  Although  diluted  alcohol  causes  shrinking,  by  em- 
ploying absolute  alcohol  (not  under  99.5  per  cent)  we  obtain  in  the 
case  of  compact  structures  (spleen,  kidneys,  digestive  glands,  etc.)  a 
fixation  without  shrinking.  The  explanation  of  this  is  found  in  the 
double  action  of  alcohol,  both  precipitating  and  chemical.  The 
latter  which  effects  the  transformation  of  sols  and  reversible  gels 
into  irreversible  gels  requires  a  certain  time  and  indeed  more  time 
the  more  dilute  the  alcohol  is,  so  that  we  must  endeavor  to  hurry 
the  chemical  action  as  much  as  possible  by  employing  concentrated 
alcohol.  The  double  action  of  alcohol  may  be  easily  demonstrated: 
If  a  solution  of  albumin  is  poured  into  alcohol  a  moderate  amount  of 
precipitate  is  formed  which  dissolves  again  upon  diluting  with  water; 
the  longer  the  time  that  elapses  before  diluting,  the  less  is  dissolved 
and  the  further  has  the  chemical  coagulation  process  advanced. 
Besides  ethyl  alcohol,  methyl  alcohol  may  be  employed. 

Formaldehyd  (Formol  or  Formalin).  The  40  per  cent  formol  solu- 
tion in  the  shops  is  usually  diluted  10  times  with  water;  if  we 
speak  of  10  per  cent  formol  solution  we  mean  that  it  contains  4 
per  cent  of  formaldehyd.  Such  a  solution  is  preferable  for  uniform 
fixation  and  preservation  of  compact  organs  (liver,  brains);  it  is 
less  desirable  for  cell  and  nuclear  structures.  A  4  per  cent  formalde- 
hyd solution  is  the  best  preservative  for  scientists  on  collecting  ex- 
peditions, even  though  it  is  not  well  adapted  for  fixation.  After 
formol  fixation  the  staining  is  often  not  all  that  could  be  desired. 
The  preeminent  properties  of  formol  depend  on  the  fact  that  it  is 
chemically  very  active,  easily  diffusible  and  hardly  at  all  adsorbed 
by  organic  substances.  The  chemical  process  of  tanning  which  is 
very  similar  to  fixing,  is  much  less  complicated  with  formol  than 
with  tannin  and  pyrogallic  acid,  with  which  an  adsorption  precedes 
the  chemical  change.  These  two  substances  are  hardly  ever  em- 
ployed alone;  at  times  they  follow  fixation  with  osmium. 

We  have  now  reviewed  the  most  important  substances  used  as 
fixatives.  In  practical  histology  almost  all  of  them  are  used  in 
mixtures.  We  employ  chromic  acid  +  acetic  acid,  potassimn  bi- 
chromate -f-  sublimate  +  glacial  acetic  acid,  nitric  acid  +  potassium 
bichromate,  osmic  acid  +  potassium  bichromate,  chromic  acid  -f- 
picric  acid  +  nitric  acid,  alcohol  +  glacial  acetic  acid,  etc.  Direc- 
tions for  fixing  tissues  are  legion,  but  they  are  "cooking  recipes" 


424  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

without  any  thought  whatsoever  of  the  chemistry  involved.  For  a 
truly  scientific  "theory  of  fixation"  it  would  be  necessary  first  to  con- 
sider the  condition  of  the  solutions  involved,  so  that  even  the  funda- 
mental facts  would  have  to  be  experimentally  determined,  and  then 
only  would  we  be  in  a  position  to  determine  their  action  on  colloids. 
At  present  we  lack  almost  the  very  essentials  for  the  development 
of  a  rational  method  from  this  wilderness  of  directions. 

Hardening. 

Hardening  follows  fixation.  For  this  purpose  alcohol  is  almost  ex- 
clusively used;  it  is  gradually  concentrated,  beginning  with  50  per 
cent  alcohol  and  then  increasing  the  strength  10  per  cent  until  96 
per  cent  alcohol  is  reached.  A  preliminary  washing  out  of  the 
fixative  is  required,  only  if  it  forms  precipitates  with  alcohol. 

In  order  to  make  thin  sections  with  the  microtome  or  razor  it  is 
usually  necessary  to  embed  the  preparation;  in  structures  contain- 
ing lime,  siliceous  or  chitinous  deposits,  these  must  be  removed  first 
by  employing  suitable  acids.  Finally,  the  sections  must  be  mounted. 
We  shall  not  discuss  these  manipulations  at  greater  length,  since 
they  are  purely  technical. 

From  our  viewpoint,  however,  a  very  important  procedure  is 

Staining. 

Unstained  specimens  are  usually  so  uniformly  transparent  that  it 
is  difficult  or  almost  impossible  to  distinguihs  their  intimate  struc- 
ture. In  order  easily  to  recognize  the  individual  structures,  histol- 
ogists  make  use  of  stains.  As  has  been  said,  they  are  chiefly 
concerned  with  a  morphological  classification;  to  see  cells,  it  usually 
suffices  to  stain  the  nuclei;  cell  division,  spermatogenesis  and  secre- 
tion require  specific  stains.  It  is  remarkable  that  but  few,  especially 
P.  Ehrlich,  P.  G.  Unna  among  others,  have  considered  what  con- 
clusions concerning  the  chemical  nature  of  the  stained  substance  may 
be  derived  from  staining.  We  are  unacquainted  with  any  conclusions 
concerning  the  physical  nature  (density)  of  tissues.  The  elaboration 
of  these  investigations  would  be  of  great  importance,  since  staining 
pictures  for  us  the  action  of  drugs,  toxins  and  disinfectants. 

The  Theory  of  Staining. 

Though  supporters  of  the  chemical  and  of  the  physical  theories  of 
dyeing  were  until  recently  actively  disputing,  there  are  mutual  con- 
cessions at  present.    We  have  recognized  that  dyeing  does  not  occur 


MICROSCOPICAL   TECHNIC  425 

in  response  to  a  fundamental  law  but  that  various  complicating 
factors  enter,  great  variations  being  possible  by  reason  of  the  variety 
of  dyes  and  fabrics. 

Otto  N.  Witt  proposed  the  theory  that  the  dye  occurred  in  the 
fiber  in  solid  solution.  This  assumption  can  only  apply  to  the  initial 
stages  as  G.  v.  Georgievics  showed,  in  experiments  on  the  absorption 
of  acids  by  wool.  The  further  absorption  of  dyes,  in  general,  corre- 
sponds to  an  adsorption.  This  is  the  result  of  quantitative  studies 
of  the  distribution  of  dyes  between  fiber  and  liquor  (the  technical 
name  for  the  dye  solution). 

We  owe  this  knowledge  to  the  researches  of  J.  R.  Appletard  and 
J.  Walker,  W.  Biltz,  H.  Freundlich  and  G.  Losev,  G.  v.  Geor- 
gievics and  L.  Pelet-Jolivet;  these  experiments  show,  moreover, 
that  there  is  no  essential  difference  between  the  adsorption  of  formic 
acid  by  blood  charcoal  and  of  indigo-carmine  by  silk.  What  ap- 
plies to  textile  fibers  we  may  apply  as  well  to  animal  and  plant 
tissues. 

In  technical  dyeing  which  has  formed  the  chief  basis  of  theoreti- 
cal studies,  the  addition  of  electrolytes  (NaCl,  Na2S04,  etc.)  are  im- 
portant factors  which  modify  the  state  of  swelling  of  the  fabric  and 
markedly  influence  the  tendency  of  the  more  or  less  colloidal  dye  to 
precipitate;  in  biological  staining  electrolytes  are  not  used  to  such 
an  extent,  but  they  always  enter  as  factors. 

The  course  of  the  adsorption  curve  requires  that  proportionately 
much  more  dye  shall  be  removed  from  a  very  dilute  solution  than 
from  one  that  is  more  concentrated,  an  observation  which  impresses 
every  one  who  investigates  dyes. 

It  must  be  assumed  if  adsorption  phenomena  are  involved,  that 
the  entire  dye  may  be  removed  by  sufficiently  prolonged  washing, 
i.e.,  that  the  process  is  reversible;  this,  as  is  well  known,  is  contrary 
to  the  facts.  The  adherents  of  the  adsorption  theory  maintain  that 
traces  of  the  strongly  adsorbed  dyes  which  can  no  longer  be  recog- 
nized in  the  dye  bath  or  wash  water  are  in  adsorption  balance  with 
the  dye  taken  up  by  the  fiber. 

In  most  cases  of  true  dyeing,  fixation  may  be  brought  about  by 
secondary  intercurrent  chemical  processes  between  fiber  and  dye. 
This  firm  union  between  fiber  and  dye  is  what  the  adherents  of  the 
chemical  theory  of  dyeing  (M.  Heidenhain,  E.  Knecht,  W.  Suida 
and  his  pupils)  chiefly  advance  in  support  of  their  theory.  They  say, 
in  general,  that  the  textile  fiber  is  a  complex  organic  substance  which 
undergoes  a  double  decomposition  with  a  dye  salt,  just  the  same  as 
any  other  salt;  the  result  of  this  double  decomposition  is  on  the  one 


426  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

hand  an  insoluble  compound  (textile  fiber-dye)  and  on  the  other  a 
soluble  compound  which  passes  into  the  bath,  e.g., 

wool  +  rosanilin  hydrochlorid  =  wool  rosanilin  base  +  ammonium  chlorid. 

The  adherents  of  the  adsorption  theory  insist  that  the  dye  salts 
are  frequently  strongly  hydrolyzed  in  solution,  so  that  there  is, 
accordingly,  only  an  adsorption  of  the  color  base  or  color  acid,  but  on 
this  account  no  chemical  double  decomposition  is  required  in  the 
staining,  inasmuch  as  fibers,  as  well  as  dye,  are  often  colloids  of 
opposite  charge  which  mutually  precipitate  each  other.  In  favor  of 
this  view  is  the  fact  that  a  dye  solution  stains  the  better,  the  more 
colloidal  it  is.  The  numerous  minute  additions  employed  in  micro- 
scopical stains  (methylene  blue  with  a  trace  of  alkali,  gentian  violet 
with  anilin  water,  etc.)  are  usually  added  for  the  purpose  of  making 
from  a  true  solution  one  less  dispersed.  P.  G.  Unna,  who  first 
recognized  this,  applied  the  characteristic  term  "incipient  precipita- 
tion" to  the  condition  of  a  staining  fluid  most  suitable  for  staining. 
Finally,  we  shall  indicate  another  point  made  by  adherents  of  the 
adsorption  theory;  that  in  those  cases  in  which  hydrolytic  cleavage 
is  not  demonstrable,  we  frequently  observe,  —  not  only  in  the  case  of 
textile  fibers  but  also  in  adsorption  by  charcoal  and  silicates,  —  that 
with  the  taking  up  of  the  dye,  a  cleavage  of  the  dye  salt  occurs 
(demonstrated  for  crystal  violet,  fuchsin,  etc.),  whereby  the  cation 
(dye  base)  goes  to  the  adsorbent  and  the  anion  goes  into  the  solution. 
(This  is  chiefly  true  of  basic  dyes.) 

Such  phenomena  were  first  demonstrated  by  J.  M.  van  Bemmelen 
in  the  adsorption  of  potassium  sulphate  by  hydrated  manganese 
dioxid;  free  sulphuric  acid  is  found  in  the  solution  while  KOH  is 
adsorbed.  Masius*  has  shown  in  the  case  of  the  very  strongly 
hydrolyzed  anilin  salts,  that  more  anilin  than  acid  is  adsorbed  by 
charcoal. 

To  explain  these  cleavage  processes,  especially  in  dyeing,  it  must  be 
assumed  that  the  easily  adsorbable  dye  ion  displaces  a  cation  K,  Na 
or  the  like,  which  is  already  present  and  adsorbed,  though  possessing 
little  capacity  for  adsorption. 

The  weak  point  in  the  argument  for  the  chemical  theory  resides 
in  the  fact  that  we  do  not  know  the  real  constitution  of  the  adsorbent, 
that  is,  the  fiber  (silk,  wool,  cotton,  etc.,),  so  that  we  do  not  know  what 
chemical  groups  are  involved  in  a  dye  combination.  H.  Bechhold 
accordingly  strove  to  solve  this  question  by  using  as  adsorbent  a 
group  of  substances  whose  composition  is  accurately  known,  namely, 
naphthalin  C10H7OH,  naphthylamin  C10H7  (NH2)  and  amidonaph- 
thol  C10H6OHNH2.     The  result  of  this  experiment  is  reproduced  on 


MICROSCOPICAL   TECH  NIC  427 

page  30  and  shows  that  acid  groups  in  the  adsorl)ent  fix  color  bases 
especially;  that  basic  groups  fix  acid  colors;  and  that  the  amphoteric 
amidonaphthol  stains  very  well  with  both  acid  and  with  basic  dyes. 

In  the  matter  of  cell  staining  not  only  is  the  chemical  nature 
of  the  cell  constituents  and  of  the  dye  involved,  but  the  physical 
prerequisites  for  the  penetration  of  the  dye  must  be  supplied. 

In  the  dyes  we  have  a  group  of  substances,  most  of  them  chemically 
well  defined,  which  exhibit  all  transitions  from  true  crystalloid  {e.g., 
methylene  blue)  to  the  highly  colloidal  hydrosols  {e.g.,  benzoazurin). 
There  is,  already,  an  extensive  literature  in  reference  to  the  colloidal 
properties  of  dyes  which  has  been  collected  in  the  work  of  L.  Pelet- 

JOLIVET.* 

The  experiments  of  R.  Höber  and  S.  Chassin  *  showed  that,  in 
general,  the  more  colloidal  a  dye  is,  the  more  difficult  is  its  absorp- 
tion by  the  kidney  epithelium  of  frogs.  Some  exceptions  are  prob- 
ably associated  with  specific  chemical  properties. 

The  idea  that  the  staining  of  a  living  plant  tissue  depends  on  the 
dispersion  of  the  dye  was  developed  into  a  comprehensive  theory  by 
RuHLAND  in  his  "ultrafilter  theory"  which  points  to  a  satisfactory 
explanation  of  the  staining  processes  in  organized  tissues.^ 

An  important  element  in  staining  capacity  seems  to  me  to  have 
been  disregarded  hitherto;  it  is  the  question  of  the  density  of  the  sub- 
stance to  be  stained  in  its  relation  to  the  diffusibility  of  the  dye.  It 
is  quite  clear  that  an  easily  diffusible  dye  will  penetrate  everywhere, 
and  that  a  dye  possessing  no  diffusibility  will  always  remain  on  the 
external  surface  of  the  tissue.  If  we  are  dealing  with  substances  of 
medium  density,  it  will  depend  upon  the  density  of  the  tissues 
whether  it  will  penetrate  at  all,  and  to  what  extent.  If  we  know 
our  dyes  from  this  point  of  view  we  will  be  in  a  position  to  draw 
conclusions  from  their  penetration  as  to  the  structure  of  the  tissue 
examined.  Some  experiments  performed  with  this  end  in  view  will 
explain  what  is  meant.^  In  solving  the  previous  question  I  em- 
ployed paper  strips  which  were  soaked  in  glacial  acetic  acid  collodion  ^ 
of  different  concentrations   (8  per  cent,  4.5  per  cent,  1.5  per  cent 

1  I  am  glad  to  learn  that  J.  Traube  and  F.  Köhler,  obviously  without  know- 
ing my  views  (see  1st  edition  of  this  book,  1912,  p.  49),  in  a  recent  pubhcation, 
1915,  likewise  point  out  the  significance  of  "the  variability  of  the  dispersing  gel 
by  added  substances,"  inasmuch  as  dyes  producing  swelling  increase  permeability 
and  dyes  causing  shrinking  diminish  it.  I  do  not  consider  satisfactorilj^  established 
the  proof  of  swelling  and  the  shrinking  properties  on  irreversible  gels  and  the  con- 
clusion drawn  from  them  since  the  experiments  of  the  author  depend  onlj'  on 
reversible  gelatin  gel. 

-  These  investigations  have  not  been  pubUshed  before. 

^  A  solution  of  collodion  in  glacial  acetic  acid. 


428  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

and  0),  gelatinized  and  then  placed  in  running  water  until  every 
trace  of  acid  was  removed.  These  strips  were  then  placed  in  0.5 
per  cent  dye  solutions  for  10  minutes  and  then  washed  in  running 
water  until  the  wash  water  was  almost  entirely  colorless.  The  re- 
sults are  shown  in  the  table  on  page  429. 

It  follows  from  these  experiments  that  the  staining  is  more  intense, 
the  denser  the  stained  substance,  in  the  case  of  easily  diffusible 
dyes,  as  aurantia,  methylene  blue  and  crystal  violet.  Conversely, 
in  those  difficultly  diffusible,  as  chrome  violet  and  benzopurpurin,  the 
intensity  of  the  staining  diminishes  with  the  density.  Obviously,  the 
particles  of  the  dye  are  largely  colloidally  distributed  and  too  large 
to  penetrate  the  pores  of  the  filter.  In  between  there  are  substances 
of  medium  particle  size  in  which  ths  staining  of  the  different  samples 
does  not  vary  much  one  way  or  the  other,  as,  e.g.,  in  the  case  of 
alizarin,  janus  red,  bismarck  brown,  etc.  An  important  factor  in 
these  experiments  is  the  time  element,  as  may  be  seen  in  the  first 
column  (8  per  cent).  The  slower  a  dye  diffuses  the  longer  the  time 
that  must  elapse  for  it  to  penetrate  a  dense  tissue  and  the  slower  the 
color  constituents  that  are  not  firmly  bound  by  the  fibers  will  diffuse 
away. 

In  this  connection  experiments  of  E.  Knoevenagel*  and  of  O. 
Eberstadt*  are  of  interest;  they  tested  samples  of  acetyl  cellulose 
swollen  to  various  degrees,  for  their  capacity  to  take  up  methylene- 
blue  solution  of  0.05  per  cent.  They  found  that  the  speed  of  ad- 
sorption was  approximately  proportionate  to  the  swelling,  so  that 
dyes  which  penetrated  greatly  swollen  acetyl  cellulose  in  a  few  min- 
utes required  months  in  the  case  of  acetyl  cellulose  that  was  not 
swollen. 

Elsewhere  we  have  discussed  whether  there  are  not  other  factors 
involved  besides  the  speed  of  diffusion  and  the  size  of  the  particles. 

The  methods  of  analysis  proposed  by  me,  which  have  not  as  yet 
been  applied  to  organized  tissues,  promise  fewer  results  in  micro- 
scopic preparations  and  microtome  sections  since  the  surfaces  to  be 
penetrated  are  so  thin  that  sufficient  differences  are  not  noticeable. 
However,  we  might  expect  new  information  in  the  case  of  coarse 
pieces  of  tissue  which  are  to  be  examined  after  they  are  sectioned. 

In  what  precedes,  the  discussion  of  the  dyeing  process  has  been 
studied  only  from  the  standpoint  of  the  chemist  and  the  physico- 
chemist.  Somewhat  independently  of  it  and  with  other  means  the 
same  discussion  will  be  carried  into  biology. 

The  dye  chemist  deals  with  the  coloring  of  a  few  fibers  of  almost 
constantly  the  same  constitution,  with  silk  and  wool,  which  dye 
easily  with  most  dyes,  and  with  cotton  and  related  fibers  which  are 


MICROSCOPICAL   TECHNIC 


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430  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

dyed  directly  only  by  certain  groups  of  dyes.  On  this  account  he 
is  able  to  obtain  almost  uniform  dyeing.  The  biologist,  on  the  con- 
trary, has  to  stain  much  more  numerous  varieties  of  tissue  and  it  is 
quite  wonderful  what  different  shadings  and  even  what  different  colors 
the  individual  tissue  elements  of  a  tissue  often  show  after  treatment 
with  a  single  stain.  If  we  consider,  moreover,  that  each  individual 
tissue  element  frequently  selects  its  own  particular  constituent  from 
a  mixture  of  stains,  thus  forming  the  brightest  pictures,  we  shall 
understand  why  the  chemical  theory  of  dyeing  is  the  most  convinc- 
ing to  biologists. 

The  following  is  a  resume  of  the  results  to  date  of  the  investiga- 
tions upon  dyeing  with  basic  and  with  acid  dyes.  By  adsorption  the 
dye  is  concentrated  upon  the  tissues,  with  which  a  fixation  may  occur 
as  the  result  of  chemical  processes. 

Hitherto  we  have  only  considered  the  so-called  substantive  dyeing, 
by  which  the  fabric  stains  directly  in  the  dye  solution  without  any 
previous  preparation  (wool,  silk).  Vegetable  fibers,  such  as  cotton, 
flax,  paper,  etc.,  take  up  very  little  color  from  most  dye  solutions 
and  do  not  hold  it  very  firmly;  they  require  a  mediator  to  chain 
the  color  to  them,  namely  a  mordant.  This  type  of  dyeing  is  known 
as  adjective  dyeing,  a  term  introduced  into  industrial  dyeing  tech- 
nology by  J.  BANCROF.T.  In  biological  staining  the  chief  mordants 
are  alum  and  ferric  oxid  salts.  The  combination  of  mordants  with 
dyes  (hemotoxylin,  hematin  and  alizarin  colors)  are  called  lakes. 
W.  BiLTZ  has  definitely  proved  that  in  addition  to  physical  adsorp- 
tion, chemical  combinations  occur  in  adjective  dyeing. 

Histologically,  staining  with  dye  mixtures  is  much  employed.  P. 
Ehrlich  found  that  if  aqueous  solutions  of  an  acid  dye,  e.g.,  acid 
fuchsin  or  orange  G,  were  mixed  with  a  basic  one,  e.g.,  methylene 
blue  or  methylene  green,  so  that  one  remained  in  excess,  then  no 
precipitate  was  formed.  From  colloidal  solutions  of  dye  mixtures 
certain  tissue  elements  remove  the  basic  and  others  the  acid  dye. 
It  is  thus  possible  to  obtain  with  one  solution  double  stains,  or  even 
triple  stains  (triacid).     (See  above.) 

According  to  the  investigations  of  0.  Teague  and  B.  H.  Buxton, *2 
acid  and  basic  dyes  precipitate  most  completely  if  they  are  mixed 
in  equimolecular  proportions.  An  excess  of  one  dye  interferes  with 
precipitation,  i.e.,  it  acts  as  a  protective  colloid,  and,  in  fact,  the  in- 
terference zones  are  wider,  the  more  colloidal  the  dye.  Especially 
important  for  the  histologist  is  the  fact  that  highly  colloidal  dye 
mixtures  are  bound  more  firmly  together  than  those  that  are  slightly 
colloidal. 

We  must  consider  very  critically,  microchemical  reactions  and 


MICROSCOPICAL   TECHNIC  431 

stains  which  arise  from  the  interaction  of  two  chemical  substances 
with  the  formation  of  an  insoluble  precipitate.  R.  Liesegang*^ 
has  called  attention  to  the  phenomena  involved  in  his  investigations 
of  Golgi's  stain.  If  a  piece  of  brain  is  placed  in  potassium  bichro- 
mate, and  after  it  is  completely  soaked  through,  it  is  then  immersed 
in  silver  nitrate,  some  of  the  ganglion  cells  in  which,  silver  Chromate 
has  been  precipitated  are  stained  reddish  brown. 

The  interior  of  the  brain  substance  is  never  thoroughly  stained, 
notwithstanding  the  fact  that  potassium  bichromate  is  present  after 
the  first  process  and  silver  nitrate  after  the  silver  bath.  The  reason 
is  as  follows:  when  the  chromatized  portion  of  brain  is  placed  in  the 
silver  nitrate  solution,  silver  Chromate  forms  in  the  outer  layers; 
the  potassium  bichromate  present  in  the  interior  diffuses  outward 
where  it  is  arrested  by  the  silver  so  that  the  interior  is  more  and  more 
depleted  of  Chromate.  The  irregularities  in  the  Golgi  staining  de- 
pend upon  similar  interferences  with  diffusion  and  nucleus  actions  of 
silver  Chromate,  on  account  of  which  only  a  portion  of  the  ganglion 
cells  are  stained.  After  staining  peripheral  nerves  with  Golgi's 
stain  we  obtain  stratifications  in  the  axis  cylinders  (Fromann's 
lines).     These  have  been  shown  to  be  artifacts. 

The  Technic  of  Staining. 

We  distinguish  staining  en  masse,  section  staining  and  vital  staining. 

In  staining  en  masse  the  entire  object  is  immersed  in  the  stain 
solution  subsequent  to  hardening.  If  this  is  soluble  in  alcohol,  it 
requires  no  special  precautions;  it  is  otherwise  with  solutions  con- 
taining alum,  in  which  case  alcohol  in  the  object  must  first  be  re- 
placed by  water. 

After  staining,  the  dye  is  washed  away  with  water  or  alcohol  until 
the  fluid  remains  colorless.  After  staining  in  aqueous  solution  the 
piece  must  be  rehardened  in  alcohol.  The  subsequent  treatment  is 
then  the  same  as  in  unstained  pieces. 

Section  staining  is  much  more  frequently  employed,  since  not  only 
details  are  brought  out  better,  but  the  staining  can  be  watched  more 
closely  and  later  counter-stains  may  be  added  intermittently.  Ac- 
cording to  the  dilution  of  the  stain  solution  and  the  length  of  time  the 
section  is  stained,  we  may  obtain  on  the  one  hand  contrasting,  or  on 
the  other  finely  shaded  pictures  with  much  more  detail. 

Vital  staining,  the  staining  of  living  tissues,  was  introduced  by  P. 
Ehrlich  and  was  apphed  by  this  investigator  in  his  classical  work 
on  "The  Oxygen  Requirements  of  the  Organism"  to  the  processes  of 
living  cells.     At  present  it  has  the  center  of  interest,  and  from  it  we 


432  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

may  expect  most  valuable  discoveries  on  the  physiology  and  pathology 
of  living  tissue  as  well  as  concerning  the  mechanism  of  the  action  of 
drugs.  E.  GoLDMANN,  R,  HÖBER  and  W.  Schuleman  have  in  recent 
years  contributed  much  concerning  the  utiHzation  and  theory  of  vital 
staining.  They  studied  healthy  and  sick  animals,  whereas  Küster 
and  RuHLAND  apphed  vital  staining  to  plants.  As  yet  vital  stain- 
ing of  bacteria  and  other  microorganisms  has  not  been  definitely 
attained  (Eisenberg).  The  stain  must  not  be  poisonous  or  the  cell 
will  die  before  it  has  the  desired  color.^  We  have  numerous  dyes  at 
present  which  fulfill  this  condition.  A  few  of  the  most  useful  are 
mentioned,  methylene  blue,  neutral  red,  toluidin  blue,  trypan  blue, 
trypan  red  and  isamin  blue.  The  studies  of  Ruhland  on  plants,  as 
well  as  those  of  Evans,  Schuleman  and  Wilborn  on  animals,  indicate 
strongly  that  the  extent  of  dispersion  of  the  dye  chiefly  determines 
its  suitability  for  vital  staining,  so  that  the  cell  behaves  like  an  utra- 
filter  (see  p.  428).  A  dye  that  is  too  diffusible  distributes  itself  too 
readily  in  all  the  organs  and  is  accordingly  quickly  excreted  by  them; 
one  that  is  highly  colloidal  remains  at  the  site  of  injection.  The 
studies  of  Ruhland  include  both  basic  and  acid  dyes  while  the 
experiments  of  Evans,  Shulemann  and  Wilborn  were  only  with 
acid  dyes. 

It  was  formerly  believed  that  only  lipoid  soluble  dyes  penetrated 
living  tissues,  but  this  view  has  not  been  sustained  (see  also  Garmus)  . 
Many  vital  stains  are  known  which  are  insoluble  in  fats.  The  col- 
loidal metals  are  included  among  these;  they  have  proven  useful 
agents  in  studying  "distribution"  in  J.  Voigt's  method  of  investiga- 
tion. This  does  not  by  any  means  imply  that  lipoid  insoluble  vital 
stains  may  not  be  especially  suitable  for  some  of  the  organs  which  are 
rich  in  hpoids.  Thus,  for  instance,  axis  cylinder  and  ganghon  cells 
of  the  nerve  substance  are  most  intensely  stained  by  methylene  blue. 
It  is  remarkable  that  the  cell  nucleus  which  stains  most  intensely 
with  basic  dyes  when  the  object  is  dead,  with  vital  staining  is  con- 
stantly colorless;   nuclear  staining  occurs  only  when  the  cell  dies. 

If  vital  stains  are  to  be  fixed,  i.e.,  made  insoluble,  ammonium 
molybdate,  sublimate,  picric  acid,  etc.,  are  employed. 

If  this  fixation  is  omitted,  the  dye  diffuses  away  after  death,  i.e., 
according  to  the  changed  condition  of  the  tissue,  physical  and 
chemical,  and  a  different  distribution  results* 

1  The  lack  of  toxicity  of  vital  stains  is  only  relative;  in  concentrated  solution 
they  are  all  poisons  and  may  be  used  only  in  extreme  dilutions.  Safranin  and 
methyl  violet,  especially,  are  quite  poisonous,  and  on  this  account  they  cannot 
be  employed  for  injections  into  the  higher  animals. 


MICROSCOPICAL  TECHNIC  433 

THE  TISSUE  ELEMENTS  IN   THEIR  RELATION   TO   FIXATIVES 

AND   DYES.  , 

With  iodin-potassium  iodicl  solution,  starch  grains  give  a  blue 
adsorption  compound  (see  p.  135). 

Glycogen  forms  with  it  a  red  adsorption  compound.  The  stain 
with  strongly  alkaline  potassium  carmine  recently  recommended  by 
Best  is  so  complicated  that  it  cannot  yet  be  interpreted. 

The  Lipoids. 

The  fixation  and  staining  of  lipoids  can  hardly  be  regarded  as 
other  than  a  colloid-chemical  question.  Fixation  is  generally  ac- 
complished with  osmic  acid  by  means  of  which  the  fat  is  simultane- 
ously blackened  and  the  acid  is  reduced  to  colloidal  metallic  osmium; 
similarly,  gold,  silver  and  palladium  salts  are  reduced  to  the  colloidal 
metal.  Of  the  true  dyes,  we  must  especially  consider  those  which  are 
very  soluble  in  fat,  though  quite  indifferent  chemically,  and  which 
are  very  slightly  adsorbed  by  the  other  constituents  of  the  cell. 
Among  these  are  Scarlet  R  (fettponceau)  and  Sudan  III.  Both  are 
amphoteric  dyes  in  which  the  basic  as  well  as  the  acid  character  is  so 
indefinite  that  they  seem  quite  indifferent  and  do  not  form  salts  with 
aqueous  caustic  soda.  Employed  in  alcoholic  solution,  staining 
results. 

Protoplasm. 

We  may  attribute  to  protoplasm  chemical  properties  similar  to 
those  of  the  albumins.  Protoplasm  may  be  amphoteric,  on  which 
account  neither  acid  nor  basic  properties  become  more  prominent. 
Consequently,  protoplasm  stains  only  faintly  with  either  basic  or 
acid  dyes,  even  though  its  water  content  is  relatively  high. 

Nucleus. 

The  chief  constituents  of  the  cell  nucleus  are  the  nucleoprofeins. 
These  are  strongly  acid  in  character;  to  them  may  be  attributed  the 
intense  staining  of  the  nucleus  with  basic  dyes,  and  to  them  the  in- 
tensely staining  constituent  of  the  nucleus  is  indebted  for  the  name 
chromatin  or  chromatic  substance  among  histologists.  The  union  Tvdth 
the  color  base  becomes  firmer  with  the  lapse  of  time,  since  in  the 
beginning  it  is  possible  to  effect  almost  complete  decolorization  A^ith 
alcohol,  whereas  when  the  dye  acts  for  a  longer  period  the  nuclei 
retain  their  intense  staining  and  only  clouds  of  color  leave.  For 
nuclear  staining  any  basic  dye  may  be  employed;  safranin,  fuchsin, 
methyl  violet,  methyl  green  and  bismarck  brown  are  recommended 
most  highly. 


434  COLLOIDS  IN  BIOLOGY  AND  MEDICINE 

Another  favorite  nuclear  staining  method  is  with  the  mordant  dyes, 
e.g.,  hematoxyhn  or  carmine.  In  this  instance,  also,  the  acid  char- 
acter of  the  nuclear  proteins  explains  the  action  of  the  dyes.  The 
nuclear  proteins  adsorb  the  mordants,  usually  colloidal  aluminium 
hydroxid  (from  alum),  and  these  form  an  insoluble  compound  with 
the  acid  hematoxylin  or  one  of  its  oxidation  compounds,  or  with 
acid  carmine. 

Finally,  we  may  mention  the  double  staining  of  Romanowsky, 
which  has  been  modified  by  G.  Giemsa.  Its  underlying  principle 
is  that  a  basic  blue  dye  (methylene  azur  or  methylene  blue)  is  mixed 
with  an  acid  dye  eosin  (see  p.  426).  At  first  the  preparation  stains 
blue  in  the  mixture;  gradually  there  occurs  a  differentiation  into 
blue  and  red  elements  or  combination  violet  shades  whereby  the 
nuclei  become  red.  For  the  present,  all  interpretations  of  this  phe- 
nomenon are  quite  hypothetical ;  it  presents  a  very  interesting  colloid- 
chemical  problem.  If  methylene  azur  and  eosin  are  mixed,  a  colloidal 
solution  of  eosin-acid-methylene  azur  forms,  provided  that  one  of  the 
two  dyes  is  present  in  excess.  Nuclear  staining  may  occur  in  such 
a  way  that  the  basic  methylene  azur  serves  as  mordant  for  the  eosin; 
it  is  also  possible  that  the  nuclei  stain  better  with  colloidal  eosin- 
acid-methylene  azur  than  with  crystalloidal  methylene  azur,  and  that  in 
a  reaction  which  requires  time  (possibly  hydrolytic  cleavage)  the  red 
color  base  of  methylene  azur  becomes  free.  In  this  double  staining 
there  enter  as  factors  phenomena  involving  the  colloidal  condition 
of  both  dye  and  specimen  with  respect  to  the  diffusibility  of  the  dye 
and  perhaps  also  other  circumstances  which  have  not  been  consid- 
ered here.  This  may  be  assumed  both  from  the  accurate  directions 
which  are  given  for  the  preparation  and  age  of  the  solution,  the 
thickness  of  the  preparation,  the  duration  of  staining,  etc.,  and 
from  the  fact  that  every  departure  from  the  directions  gives  a  dif- 
ferent result. 

Connective  Tissue,  Capillary  Walls,  Membranes,  Etc. 

From  the  numerous  reports  I  gather  that  only  easily  diffusible 
stains,  especially  the  sulphoacids  (acid  fuchsin,  soluble  blue  com- 
bined with  picric  acid),  are  suitable  for  this  purpose.  This  probably 
depends  upon  the  fact  that  connective  tissue,  etc.,  are  among  the 
tissues  poorest  in  water  and  least  swollen,  so  that  dyes  of  more  col- 
loidal character  are  unable  to  penetrate  them. 

For  the  staining  of  elastic  fibers,  which  is  best  performed  by  the 
orcein  method  of  P.  G.  Unna  and  Taenzer  or  by  Weigert's  method, 
we  have  no  explanation  whatever.  The  recent  investigations  of  the 
keratins  by  L.  Golodetz  and  P.  G.  Unna  show  that  we  are  dealing 


MICROSCOPICAL   TECHNIC  435 

with  a  number  of  chemically  very  different  substances  (ovokeratin, 
neurokeratin,  elastin).  [Van  Gibson's  stain  contains  picric  acid  and 
stains  elastin  specifically.     Tr.] 

The  Staining  of  Bacteria. 

Most  cocci  and  bacteria  have  a  definite  acid  character  evidenced 
by  the  fact  that  they  migrate  to  the  anode  in  the  electrical  current 
(see  p.  205). 

Though  they  usually  stain  intensely  with  basic  dyes  (f uchsin,  meth- 
ylene blue,  thionin,  etc.),  nevertheless  bacteria  exhibit  considerable 
differences  in  staining  capacity.  Though  all  cocci  with  which  I  am 
acquainted  stain  very  intensely,  some  bacteria,  e.g.,  paratyphoid  and 
bacilli  of  hog  erysipelas,  are  stained  more  faintly.  Spores  stain  with 
especial  difficulty,  the  more  poorly  the  older  they  are;  it  is  obvious 
that  the  solid  capsule  offers  great  resistance  to  the  penetration  of 
the  stain.  The  tubercle  bacillus  is  most  difficult  to  stain,  which  may 
be  attributed  chiefly  to  its  high  keratin  content,  inasmuch  as  other 
keratin-containing  substances  (bristles,  hair,  epidermis,  etc.)  stain 
just"  as  poorly.  The  difficulty  in  staining  the  tubercle  bacillus  was 
formerly  attributed  to  the  wax  contained.  Helbig,  however, 
showed  that  complete  removal  of  the  wax  did  not  increase  the  stain- 
ing capacity. 

Gram's  stain  is  quite  unique;  it  is  extensively  employed  for  the 
classification  of  bacteria  (we  distinguish  Gram-positive  and  Gram- 
negative).  It  is  performed  as  follows:  we  first  stain  with  methyl 
violet  or  some  related  basic  dye  and  then  subject  the  specimen  to  the 
action  of  iodin  (dissolved  in  KI).  After  this  treatment,  some  bac- 
teria readily  give  up  the  dye  to  alcohol  and  are  decolorized,  whereas 
others  firmly  retain  it.  In  the  latter  case,  a  firm  combination  has 
been  formed.  A  thorough  study  from  modern  points  of  view  would 
be  of  great  value,  since  it  would  explain  the  difference  in  the  nature 
of  the  two  groups  of  bacteria.  It  is  important  to  mention  that,  by 
Gram's  method,  a  differentiation  of  the  structm-e  of  individual  bac- 
teria may  be  revealed.  The  so-called  Babe's  corpuscles  are  not  de- 
colorized by  a  brief  action  of  alcohol.  Upon  this  fact  depends  M. 
Neisser's  method  for  identifying  diphtheria  bacilli. 

We  have  as  yet  little  insight  into  the  actual  basis  of  differentiation 
by  Gram's  stain.  It  has  actually  only  been  established  that  Gram 
positive  bacteria  show  a  greater  permeability  for  dies,  stain  more 
quickly  and  intensely  and  retain  the  dye  more  strongly  upon  de- 
colorization  with  alcohol.  Probably  the  onlj^  purpose  of  the  treat- 
ment mth  iodin  is  to  increase  the  size  of  the  dye  molecule  or  increase 
its  fixation  by  the  bacillus  (Eisenberg)  . 


> 


AUTHORS'   INDEX 


^^  refers  to  footnote  in  text 
*  refers  to  reference  given 


Abbe,  124,  127 

Abderhalden,  E.,  32,  34,  72*^,  92,  187, 
210,  210^»,  220*1,  321 

1)  Lehrbuch  d.  physiol.  Chemie  (Ber- 
hn,  1906) 

2)  Zeitschr.  f .  physiol.  Chemie  37,  484 
(1903) 

and  Guggenheim,  M.,  34*,  189* 

Zeitschr.  f.  physiol.  Chemie  54  (1908) 

and  Pettibone,  185* 

and  Strauch,  F.  W.,  187 

Zeitschr.  f.  physiol.  Chemie  71,  315- 
318  (1911) 
Abramow,  S.,  206 
AcHARD  and  Weill,  E.,  371 
Adam,  P.,  311 
Adie,  57* 

Journ.  Chem.  Soc.  344  (1891) 
Adler,  H.  M.,  363 

Journ.  of  Am.  Assoc.  2,  752  (1908) 

and  Herzog,  B.,  27* 

and  Ringer,  379 
Aggazzotti,  a.  and  Foa,  C,  374*,  384* 
Albarran,  340 
Albrecht,  E.,  305* 

Verh.  d.  D.  pathol.  Ges.  5,  2  (1903) 
Albü,    a.    and   Neuberg,    C,    218*, 
219*,  233* 

Physiol,  u.  Pathol,  d.  Mineralstoff- 
wechsels (BerHn,  1906) 
Alexander,  Jerome,  42-''",  71,  174,  188, 
195^" 

and  Bullowa,  J.  G.  M.,  174,  349* 

Archives  of  Pediatrics  (N.  Y.,  Jan., 
1910) 

(and  Zsigmondy),  247 
Altmann  and  Sachs,  H.,  206,  208 
Amann,  J.,  343* 

BuU.  Soc.  Vaudoise,  38,  131 
Ambard,  336 
Amberger,  C,  11,  366 


Ambronn,  H.,  259* 

KolL  Zeitschr.  6,  222-225  (1910) 
Ames  and  Bauer,  J.,  231,  252 
Anderson,  10 

Ariens  and  Kappers,  C.  U.,  261* 
Folia    Neuro-Biologica    1,   No.   244 
(1908) 
Araki,  F.  and  Zillessen,  H.,  226 
Arrhenius,   Sv.,   26*,   53,   56*,   105*, 
196,  197,  200*,  213,  386 
Immunochemie  (Leipzig,  1907) 
and  Herzog,  R.  O.,  47 
Arric,  Le  Fevre  de,  371,  372 
AscHERSON,  35*,  347,  347* 

Arch.    f.    Anatom,    u.    Physiol.    53 
(1840) 
AscoLi,  M.,  109,  211,  371*1,  372* 
Koll.  Zeitschr.  5,  186  (1909);  6,  293- 

298  (1910) 
and  Izar,  G.,   211,   365*,   366,   371, 
373*,  377*,  383^" 

1)  Berliner  Klin.  Wochenschr.  4  and 
21  (1907) 

2)  Biochem.  Zeitschr.  5,  394;  6,  192; 
7,  143;  10,  356;  14,  491;  17,  361 
(1907-1909) 

3)  BoU.  della  Soc.  Med.  Chir.  di 
Pavia  35  (1908) 

*)  Comptes    rend,    de    la    Soc.     de 
Biologie  65,  59  and  426 
AsHER,  L.,  324* 

Biochem.  Zeitschr.  14  (1908) 
AuER,  413 

Amer.  Journ.  of  Physiol.  17  (1906) 
and  Journ.  of  Biol.  Chem.  4  (1908) 
Auerbach  and  Pick,  330 

Babcock,  S.  M.  and  Russel,  H.  L., 
175* 
Ber.    Landw.    Ver.    Stat.    d.    Univ. 
Wisconsin   (Ver.  St.  v.  N.  A.)   20 


437 


438 


AUTHORS'   INDEX 


Babe,  435 
Bachman,  9*,  10 

Bachmann,   L.   and  Rünnstrom,   J., 
265,  265* 
Biochem.  Zeitschr.  22,  290  (1909) 
Balyer,  a.  von,  27 
Baisch  and  Landwehr,  343 
Bancroft,  J.,  413*,  430 
W.  D.,  37,  40,  46 

W.    D.    and    Clowes,    G.    H.    A., 
36-^" 
Bang,  J.,  139,  244* 

Biochem.  Zeitschr.  16,  255  (1909) 
Barbieri  and  Carbone,  352 
Barcroft,  J.  310 

1)  Journ.  of  biol.  Chemistry  3,  191 
(1907);  Pflüger's  Arch.   122,   616 
(1908) 
and  Brodie,  334 

Journ.  of  Physiol.  32,  18,  33,  52 
Bary,  de  and  Stahl,  E.,  286 
Battelli  and  Stern,  386 
Bauer,  J.,  231,  252 

and  Ames,  231,  252 
Bayliss,  W.  M.,  29,  137,  166,  182,  183, 
365 
and  Fischer,  M.  H.,  220^" 
Bechhold,  H.,  6,  10,  11,  16,  17,  26,  29 
35,  36*1,  42^  55*2^  55,  58*^,  80,  83 
84*1, 86, 95, 95*S  95^",  96^  "-*S  96^"- 
97,  97^"-,  98,  99,  99-^"-*«,  100-^"-*« 
102,  102*S  102*^  104*2,  106,  108 
115,  119,  119*"-,  125,  135*S  138 
144,  146,  147/",  148,  157*,  158** 
164,  165,  166,  180*8,  196,  197*^ 
198*4,  199*4^  201*4,  203,  203*i 
205*1,  205*1°,  241,  260,  262,  262*2 
268*,  283*,  288*,  327,  332*1^,  332*" 
333,  347*1,  348*4,  352/»,  371 
382*1,  394*^  395-''",  396*^,  401*» 
402*3,  404 

1)  Zeitschr.  f.  physikal.  Chemie  48 
385-423  (1904) 

2)  Zeitschr.  f.  physikal.  Chemie  52 
185-199  (1905) 

3)  Wienerklin.  Wochenschr.  1905,  No. 
25 

4)  Zeitschr.  f.  physikal.  Chemie  60, 
257-318  (1907) 

^)  Münchner  med.  Wochenschr.  1908, 
No.  34 


Bechhold,  H.,  ^)  Zeitschr.  f.  physikal 

Chemie  64,  328-342  (1908) 
^)   Zeitschr.  f.  physiol.   Chemie  52, 

177-180  (1907) 
8)  Koll.  Zeitschr.  2,   Nos.   1  and  2 

(1907) 
^)  Zeitschr.  f.  Hygiene  u.  Infektion- 

skrankh.  64,  113-142  (1909) 
1")  Münchner      med.      Wochenschr. 

1907,  No.  39 

11)  Koll.  Zeitschr.  5,  22-25  (1909) 

12)  van  Bemmelen-Festschrift  (1910) 
and  Ehrlich,  P.,  394*,  396,  404,  408 
Zeitschr.  f.  physiol.  Chemie  47,  173- 

199  (1906) 
and  Ziegler,  10*,  35,  54*^,  55,  55*2, 
57*1,  73^  106,  138*2,  147  /«^  143*3^ 
162*2,  238*1,  243,  244,  260*i,  269*% 
319,  320*2,  327,  329,  343,  378*% 
380,  403*2 

1)  Ann.  d.  Phys.  (4)  20,  900-918  (1906) 

2)  Zeitschr.  f.  physikal.  Chemie  56, 
105-121  (1906) 

3)  Biochem.    Zeitschr.    20,    189-214 
(1909) 

4)  Berliner  klin.  Wochenscher.  (1910) 
Beck  and  Hirch,  113 

Beckmann,  E.,  115 
Behring,  E.  von,  144 

Beitr.  z.  exper.  Therapie  10  (1905) 
Beijerinck,  M.  W.,  76 
Bemmelen,  J.  M.  VAN,  9,  26,  67,  152-''", 

217,  329,  337 
Bence,  J.  and  Koränyi,  A,  von,  315 
Benedicenti  and  Revello-Alves,  157 
Benedict,  H.,  312* 

Pflüger's  Arch.  115,  106  (1906) 
Beniasch  and  Michaelis,  204 
Bentner,  G.,  230 

R.  and  Loeb,  J.,  240,  295,  295^" 
Berczeller,  143* 

and  Czäki,  29* 
Berg,  W.  and  Fischer,  A.,  418 
Berghaus,  W.,  359 
Berman,  17 
Bernoulli,  E.,  381 
Bernstein,  296 

E.  P.  and  Simons,  Irving  E.,  211^'' 
Berthold,  G.,  283* 

Studien  über  Protoplasmamechanik 
(Leipzig,  1886) 


AUTHORS'   INDEX 


439 


Bertholet,     M.    and    Jungfleisch, 

20 
Bertrand,  G.,  191 
Best,  433 
Betegh,  von,  102 
Bezold,  a.  von,  265 
BiERRY  and  Henri,  V.,  191* 

Comptes  rend,  de  la  Soc.  de  Biologie 

60,  479  (1906) 
Billiter,  J.,  86* 

Zeitschr.  f.  physikal.  Chemie  51,  142 

(1905) 
BiLTz,  W.,  26*1,  28,  31*S  47,  86,  106, 

107,    110,    125,    135,    135*,    145*^ 

196,  197,  200*3,  384*2,  430 

2)  Ber.  d.  deutch  chem.  Ges.  37,  3138 
(1904) 

3)  Zeitschr.   f.   Elektrochemie,    1904, 
No.  51 

')  Biochem.  Zeitschr.  23  (1909) 
and  Freundlich,  H.,  110 
and  Gatin-Gruszewska.  L.,  73 
Pflüger's  Arch.  105,  115  (1904) 
Much,  H.  and  Siebert,  C,  196,  198*, 

199*,  204* 
in  E.  von  Behring's  Beitr.  z.  exper. 

Therapie  10 
and  Steiner,  H.  28* 
Koll  Zeitschr.  7,  113  (1910) 
and  Vezesack,  A.  von  46,  73*,  106*, 

106  '"•,  237*  290* 
Zeitschr.  f.  physikal.  Chemie  66,  68, 
357-382  (1909);  73,  481-512 
Bischoff,  E.,  215^",  218*,  219* 

Zeitschr.     f.     ration.     Medizin    20, 
75-118  (1863). 
Blasel  and  Matuta,  J.,  156''^'' 
Blom,  184* 
Bloor,  120 
Blunschly,  H.,  310 
BoBERTAG,    O.    and    Feist    Fischer, 
H.  W.,  67* 

BODENSTEIN,  M.,  187 

and  Dietz,  187 
BöHi,  55* 
Bohr,  Chr.,  309 

BOKORNY,  383 

BoNDi,  S.  and  Neumann,  A.,  247,  247*, 
248-'"",  373 
Wiener  klin.  Wochenschr.  20  (1910) 


BORÜET,  J.,  207 

•     Ann.  de  I'Inst.   Pasteur,  1899,  225; 
1900,  257;  1903,  161. 
and  Gengou,  207 
and  Massard,  2SG 
BoRKowsKi  and  Dunin,  205* 
BoROWiKOW,  267,  267*,  278,  279 
BosANYi  and  Mansfeld,  386 
Bosworth,  323 

BoTTAZzi,  F.  and  D'Errxco,  G.,  136* 
Pflüger's  Arch.  115,  359  (1906) 
P.  164,  165,  165*1,  228,  294 
Rend.  d.  R.  Acad.  d.  Line.  17,  ser. 
5"    (1908);    2,    sem.   fasc.    9,    10; 
18,  ser.  5"  (1909);    2,  sem.  fasc. 
8,  9, 10;  19,  ser.  5"  (1910);  2,  sem. 
fasc.  4,  210 
Ph.  and  Onorato,  340* 
Arch,  di  Fisiol.  1,  3  (1904) 
BoTTERi,  A.  and  Landsteiner,  K.,  199* 
BOURGUIGNON,  373* 

Compt.  rend.   Soc.  Biol.   64  M  22, 
1090 
BoviE,  144 
Boyle,  51 

and  Gay-Lussac,  51 
Bredig,  G.,  9, 83,  99, 154*,  184, 187,  366, 
371,  373,  374 

1)  Zeitschr.  f.  physikal.   Chemie  51 
(1898) 

2)  Zeitschr.   f.   Elektrochemie  6.    33 
(1899). 

and  Fajans,  188* 

Zeitschr.  f.  physikal.  Chemie  73,  25 
(1910) 

and  Fiske,  188 

and  Svedberg,  Th.,  4 
Brieger,  E.  and  Fischer,  H.  W.,  309 
Brodie  and  Barcroft,  J.,  334 
Brown,  Robert,  49,  49-'^" 
Brownian-Zsigmondy  Movement,  53, 

83 
Bruns,  J.  and  Spiro,  K.,  401*,  403* 
Bruyn,  L.  de,  41-'^" 
bubanovic,  386 
Büchner,  98 

G.  and  Klatte,  184* 
Bugarszky,  J.,  Roth  and  Steyrer,  336 

and  Taugl,  K.,  336 

St.  and  Liebermann,  L.,  152*,  153* 

Arch,  f .  d.  ges.  Physiol.  72,  51  (1898) 


440 


AUTHORS'   INDEX 


BuGLiA,  G.,  296 

BuLLowA,  J.  G.  M.,  169^",  174,  175, 
234 
See  also  Translator 
and  Alexander,  J.,  174,  349* 

BuNSEN,  R.,  363,  384 

BuRiAN,  R.,  59*1,  102*1,  102*2,  333 

1)  Arch,   di  Fisiol.  7   (1909)    (Fano- 
Festsch.) 

2)  Pflliger's   Arch.    d.    Physiol.    136, 
741-760 

BuRRi,  126 

BuRRiDGE,   W.,   234,   292,   298,   298^, 

298^,  325,  354,  361 
Burton,  R.,  84 
BÜTSCHLI,  O.,  10,  67 
Buxton,  B.  H.,  84*^ 
and  Rahe,  A.  H.,  203* 
Journ.  of  med.  Research  20,  No.  2 

(1909) 
Shaffer,  P.  and  Teague,  O.,  203* 
Zeitschr.    f.    physikal.    Chemie    57, 
47-89  (1907) 
and  Teague,  O.,  84*^,  430*^ 

Calcar,    Van    and    Bruyn,    L.    de, 
41/n 

Camerer,  Jr.,  265 
Carbone  and  Barbieri,  352 
Carrel,  Alexis  and  Burrows,  246* 

1)  Journ.  of  Exper.  Med.  (1910-1911) 

2)  Berliner  Idin.  Wochenschr.  (1911, 
No.  30) 

and  Dehelly,  404,  409 
Cavadias,  J.,  375 
Cervello  and  Le  Monaco,  362 
Chabonier,  336 
Chalupecky,  144 
Chamberland,  187,  190 
Charrin,   Henri,   V.  and  Monnier- 

VlNNARD,   374* 

Chassin,  S.  and  Höner,  R.,  427* 
Chiari,  R.,  68,  161 

R.  and  Januschke,  222 
Chick,  H.  and  Martin,  C.  J.,   143*, 
146* 

Journ.  of  Physiol.  40,  404  (1910) 
Chirie  and  Monnier-Vinnard,  374* 

Comptes  rend,  de  la  Soc.  de  Biologie 
61,  673  (1906) 
Clausius,  50 


Clowes,  G.  F.  L.,  109,  240,  241,  242,  378 
G.  H.  A.,  12,  36^",  37^",  38,  40 
and  Bancroft,  W.  D.,  36-''" 
and  Fischer,  M.  H.,  12,  175 

Coehn,  a.,  78* 
Ann.  d.  Physik  64,  217  (1898);   30, 
777  (1909) 

CoNHEiM,  Julius,  223 
Otto,  320 

CoRi,  J.,  417 

CORNALBA,  G.,  346 

Rev.  zen.  d.  Lait  7,  33,  56  (1908) 

Cramer,  345 

Crede,  365,  366 

1)  Apotheker-Zeitung  11,  165 

2)  Kongress    d.    D.    Ges.    f.    Chem. 
Ther.,  Oct. 

")  Berliner  klin.  Wochenschr.  No.  37 

(1901) 
4)  Arch.  f.  klin.  Chirur.  No.  69  (1903) 
^)  Zeitschr.  f.  arztl.  Fortbildung  No. 
20  (1904) 
CusHNEY,  A.  R.,  336 

and  Wallace,  J.  B.,  319* 
CzAKi  and  Berczeller,  29* 
Czapek,  F.,  240,  248 

1)  Ber.  d.  D.  Botan.  Ges.  28,  159-169 

(1910) 
and  Traube,  J.,  240  386 

Dabrowski,  72,  103,  105^"-,  146* 
Dakin,  405 

and  Dunham,  405 
Dam,  H.  van,  164* 

Chemisch  Weekblad  7,  1013-1019 
Davenport,  264* 

Boston   Soc.    Nat.   Hist.   28,    73-84 
(1877). 
Davidson  and  Michaelis,  202 
Davis,  J.,  31 
Dean,  207 
Dekhuyser,  C.  and  Stoeltzner,  W., 

420 
Delaunay,  365 
Demoor,  J.,  335 

and  Philippson,  298* 

Bull,   de  I'acad.   de  med.  de  Belg., 
655  (1908-1909) 
Denham,  W.  S.,  187* 

Zeitschr.    f.    physikal.    Chemie    72, 
641-694  (1910) 


AUTHORS'   INDEX 


441 


Denys,  G.  and  Leclef,  288 
Determann,  H.  a.,  113,  114,  114^"-. 
310  310*,  311 

Medizin.  Klinik  No.  27  (1910) 
Determeyer  and  Wagner,  343* 

Biochem.  Zeitschrift,  7,  388  (1908) 
Devaux,  34 
DiETZ,  187 
Dioscorides,  364 
Dender,  309 
Donnan  and  Harris,  46 

F.  G.,  59,  59*,  61,  62 

and  Donnan,  W.  D.,  343*,  344* 

S.,  47 

W.  D.  and  Donnan,  F.  G.,  343*,  344* 

Brit.  Med.  Journ.,  Dec.  23  (1905) 
Dreyer,  G.  and  Hausen,  144 

and  Sholto,  J.,  28* 

Proc.   of  Roy.   Soc.  82B,   168,   185 
(1910) 

Sholto,  J.  and  Douglas,  C.,  200 
DucLAUX,  J.,  91,  95,  102 

and  Malfitano,  G.,  92,  102 
Düngern,  E.  von,  202* 

Zentral,  bl.  f.  Bakt.  34,  4  (1903) 
Dunin  and  Borkowski,  J.,  205* 

Anz.  d.  Akad.  d.  W.  Zsepsch-Krakan, 
No.  7B,  608  (1910) 
DupRE,  Fr.  and  Hermann,  348* 
Durham  and  Gruber,  202-'^" 
DuRiG,  A.,  216,  289 

Pfliiger's  Arch.  85,  401-504  (1901) 

Ebbecke,  W.,  342* 

Biochem.  Zeitschr.  12,  485  (1908) 
Eberstadt,  O.,  428* 

Diss.  Heidelberg  (1909) 
Ebler,  88 

Edinger,  L.  and  Mayer,  421 
Effront,  Jean,  182 
Ehrlich,  P.,  31,  195*,  197,  201,  246, 
251,  360,  361,  430 
Ivlin,  Jahrb.  6  (1897) 
Münchner  med.  Wochenschr.  Nos. 

33,  34  (1903) 
Deutsch,     med.     Wochenschr.     597 

(1898) 
and  Bechhold,   H.,  394*,   396,   404, 

408 
and  Unna,  P.  G.,  424 
Einstein,  A.,  50,  54 


Eisenberg,  200,  201,  307,  402,  435 
and  Landst(;in(>r  and  Volk,  201 
and  Okolska,  403 
and  Volk,  200,  200*,  203 
Zeitschr.  f.  Hygiene  40,  155  (1902) 

ElSSLER,  135 

and  Pring.sheim,  135 
Ekroth,  Clarence  V.,  169''""' 
Elias,  208 

Ellenger  and  Spiro,  299 
Ellis,  Risdale,  87 
Elsberg,  C.  A.,  261 
Elsner,  Fr.,  175 
Emslander,  f.,  179*3,  179*,  180,  180*', 

181*2 

1)  Ivoll.  Zeitschr.  5,  25  (1909) 

2)  Koll.  Zeitschr.  6,  156  (1910) 

3)  Koll.  Zeitschr.  7,  177  (1910) 
and  Freundlich,  180* 

Zeitschr.  f.  physilval.  Chemie  49,  322 
(1904) 

Rieh,  179-^",  180* 
Engelmann,  Th.  W.,  296 
Engels,  218*,  220 

Arch.  f.  exper.  Pathol,  u.  Pharmakol. 
51,  346-360  (1904) 
Epstein.  A.  A.,  339 
Errico,  G.  d',  136*,  330 

and  Jappelli,  296 

and  Savarese,  330 
Etienne,  G.,  375* 

Rev.  med.  de  l'Est  1  (Sept.,  1907) 
Euler,  H.,  53,  182 
Ewald,  R.  and  Strassburg,  226 

Faraday^,  Michael,  75,  125 

Feist,    C.    and    Bobertag,    O.    and 

Fischer,  H.  W.,  67* 
Fellner,  88 

Field,  C.  N.  and  Teague,  O.,  205* 
Journ.  of  Exper.  Med.  9,  No.  1,  pp. 
86-92 
FiLEHNE,  W.,  324* 
Berliner  klin.Wochenschr.  No.  3(1898) 
Arch,  inter,  de  Pharmacodynamic  F, 
No.  133  (1900) 
FiLippi,  E.,  371,  372 

Lo  sperimentale  62,  503-522  (1908) 
Arch,  italicnnes  de  biologie  50,  175- 

189  (1908);  51,  447-456  (1909) 
and  Rodolico,  372 


442 


AUTHORS'   INDEX 


FiNDLAY,  A.,  135*,  147* 

Koll.  Zeitschr.  3,  169  (1908) 

FiNKELSTEIN,  H.,  323 

Fischer,  17,  37 
A.,  418,  421,  422 
and  Berg,  Waltlier,  418 
Emil,  5,  133,  182 
H.  W.,  216*1,  384*2^  385 

1)  Beitr.  z.  Biol.  d.  Pflauzer  (1910) 

2)  Biochem.  Zeitschr.  27,  223-245 
(1910) 

Bobertag,  O.  and  Feist,  C,  67* 

Ber.  d.  D.  Chem.  Ges.  3675  (1908) 

and  Brieger,  E.,  309 

and  Jensen,  P.,  222*,  292 

Biochem.  Zeitschr.  20, 143-165  (1909) 

Martin  H.,  12,  33,  36,  36^",  67*,  68, 
68^'^-,  70,  115,  160*,  175^",  224^"-, 
225,  226*,  227,  228*2,  229'"-,  229, 
230,  230*1,  231,  232,  233,  236,  267, 
282,  289,  289*,  290,  306*^",  315, 
316,  320*,  321,  321^",  323*,  323^''. 
324,  327,  328^",  332^",  333*,  334, 
336,  338,  338*1,  339,  344*i,  352. 
410,  411 

Das  Oedem.  in  exper.  v.  Therapeut. 
Unterricht  d.  Physiol,  u.  Pathol  d. 
Wasserb.  im.  Organismus  (Dres- 
den, 1910) 

Die  Nephritis,  eine  exper.  u.  kritische 
Studie  über  Natur  u.  Ursachen, 
d.  Prinz,  ihrer  Bechandlung  (Dres- 
den, 1911) 

1)  Kolloidchem.     Beihefte.    2,    304. 
(1911) 

2)  Koll.  Zeitschr.  8,  159-167  (1911) 
and  Bayliss,  220^« 

and  Clowes,  12,  175 

and  Henderson,  L.  J.,  232^" 

and  Hogan,  220^'^ 

and  Hooker,  M.  O.,  33,  36,  36^^  352 

and  Jensen,  P.,  291* 

and  Losev,  G.,  27* 

and  Streitmann,  A.,  296 

and  Sykes,  A.,  334 

and  Woodyatt,  335 
FiSKE,  188 
Fleischmann,  208 

Fletcher,  W.  M.  and  Hopkiijs,  F.  G., 
298 

and  Langley,  328 


Flxjri,  M.,  245* 

Flora  99,  81-126  (1908) 
FoA,    C.    and  Aggazzotti,    A.,    374*, 
384* 
Biochem.  Zeitschr.  19  (1909) 
Fornet,  A.,  178 

FouARDi,  E.,  102*,  107,  133*,  134, 134*, 
180 
KoU.-Zeitschr.  4,  185  (1909) 
L'Etat  colloidal  de  l'Amidon  (Laval, 
1911) 
Fränkel,     S.     and     Hamburg,     M., 
190* 
Beitrag  z.  chem.  Physiol,  u.  Pathol. 
8,  389-398  (1906) 
Frank,  E.  235* 

Zeitschr.  f.  physiol.  Chemie  70,  129- 
142  (1910) 
Fränkel,  S.,  183 
Frankl,  413 

Arch.  f.  exper.  Path.  u.  Pharm.  57 
(1907) 
Frei,  W.,  311* 

Zeitschr.  f.  Infektionskrank,  u.  Hyg. 
d.  Haustiere  6,  363-373,  446-475 
(1909) 
Margadant,  396,  403 
Freundlich,  H.,  14,  23,  23*,  23^"-,  24, 
24^"-,  25,  25^",  28*3,  29,  66,  78"'-. 
79,  80,  84,  87,  110,  147^",  187,  360,, 
394*1 

1)  Habilitationsschrift  (Leipzig, 
1906) 

2)  KoU.-Zeitschr.  1,   321   (1907);  7, 
193  (1910) 

(and  Emslander),  180* 

and  Losev,  G.,  27 

Zeitschr.    f.    physikal.    Chemie    59» 
284-312  (1907) 

and  Michaelis,  L.,  79 

and  Ostwald,  Wo.,  147-^" 

and  Schucht,  25* 

(and  Straub),  243 
Frey,  E.,  334*,  338,  410* 

Pflüger's  Arch.  120,  66-136  (1907) 
Friedberger,  209,  210,  371 

and  Isuneoka,  365 
Friedemann,  U.,  149,  149*,  156*i,  157, 
202-^",  207,  323*2 

1)  Arch.    f.    Hygiene    55,    361-389 
(1906) 


AUTHORS'   INDEX 


443 


Friedemann,  TJ.,  -)  in  Oppenheimer's 

Handb.  cl.  Bicdiemie  3,  2 
and  Friedenthal,  H.,  149*,  160*,  202* 
Zeitschr.  f .  exper.  Pathol,  u.  Therapie 

3,  73-88  (1906) 
and  Neisser,  M.,  84*,  86,  203,  203*, 

205*,  283* 
Friedenthal,  H.,  6,  135*i,  149*,  157*, 

164* 
Ber.  d.  D.  Chem.  Gcs.  44,  906  (1911) 
1)  Physiol.  Zeutralbl.  12,  819  (1899) 
and    Friedemann,    U.,    149*,    160*, 

202* 
Friedländer,  J.,  64*,  136* 

Zeitschr.  f.  physikal.  Chemie  38,  430 

(1901) 

2FÜRTH,  O.  VON,  169 

and  Schütz,  J.,  191* 

Beitr.  z.  chem.  Physiol,  u.  Pathol.  9, 

28-49  (1907) 
and  Lenk,  291 

Gabritschewsky,  a.,  286 
•Gaidukow,  N.,  127,  277,  278 

DmikeLfeldbeleuchtung  u.   Ultrami- 
kroskopie in  d.  Biologien.  Medezin 
(Jena,  1910) 
Galecki  and  Zsigmondy  and  Wilke- 

Dörfürt,  98 
Galitzer,  S.,  204 
Galeotti,  a.,  340* 

Arch.  f.  Anat.  u.  Physiol.  Abt.  200 
(1902) 
Gartner,  A.,  172 
-Gatin-Gruszewska,  Z.,  136* 

Pflüger's  Arch.  103  (1904) 
Gay-Lussac,  51 

and  Boyle,  51 

and  Southard,  377 
Gebhardt,  W.,  263,  269 
Gengou,  O.,  371 

and  Bordet,  207 
Gerhartz,  H.,  216*,  265,  265*i,  266 

Pflüger's  Arch.  133,  397-499  (1910) 

1)  Pflüger's  Arch.  135,  104-170  (1910) 
Gerloff  and  Kihiitz,  263'^"" 
Gesell,  R.  A.,  333 
GiBBS,  W.,  25,  25^'' 
GiEMSA,  G.,  434 
Gieson,  van,  435 
Gilbert  and  Lawes,  265 


Girard,  59*,  322,  322* 

Comptes  rend,  de  i'ac.  d.  Lc.  146, 
927  (1908);  148,  1047-1186  (1909j; 
150,  1446  (1910) 
Journ.  d.  physiol.  and  pathol.  Gen. 

12,  471  (1910) 
-Mangin  and  Henri,  V.,  204* 
Glauber,  8 
Godlewski,  88* 

Goldschmidt,  R.  and  Pribram,  387* 
Zeitschr.  f .  exper.  Pathol,  u.  Therapie 
6,  1  (1909) 
GoLGi,  264 
GoLL,  332 
Golodetz,  L.,  163 
GoLODETZ,  L.  and  Unna,  P.  G.,  434 
GoM,  W.,  366 
GoocH,  98 
Goppelsröder,  108 
Gottlieb,  R.  and  Magntjs,  R.,  332, 
336* 
Arch.  f.  exper.  Pathol,  u.  Pharm.  45, 

223  (1901) 
and  Meyer,  H.,  332*,  380*,  412* 
Graham,  Thomas,  3*,  4,  10*,  55,  84,  90, 
146 
Philos.  Transactions  1,  183  (1861) 
Liebig's  Aim.  d.  Chemie  121  (1862) 
and  Herzog,  R.  O.,  146*6 
and  Stephan,  54 
Gram,  30,  435 
Grawitz,  B.,  418^",  421 
Gros,  O.  and  O'Connor,  J.  M.,  365*, 
372* 
Arch.  f.  exper.  Pathol,  u.  Pharm.  64, 
456-467  (1911) 
Gregor,  298 

Pflüger's  Arch.,  101  (1904) 
Grosser,  O.,  102,  174,  350 
Grüber  and  Durham,  202-'^" 

and  Widal,  202^» 
Grübler,  186 
Grünwald,  411 
Grützner,  P.  von,  353 

GUAGLIERIELLO,   143* 

GuERRiNi  and  Neisser,  M.,  288* 
Guggenheim,    M.    and    Abderhalden, 

34*,  189* 
Gully,  Baumann,  329 
GUMILEVSKY,  G.  O.,  318* 

Pflüger's  Arch.  39,  566  (1SS6) 


444 


AUTHORS'   INDEX 


GÜRBER,  A.,  245,  307,  314 

and  Limbeck,  V.  and   Hamburger, 
H.  J.,  321 

Haak,  a.,  124^"-,  125 
Haas,  A.  R.  C,  388 
Haber,  f.;  59* 

Ann.  d.  Phys.  (4)  26,  927  (1908) 
Zeitschr.  f.  physikal.  Chemie  67,  385 
(1909) 
HÄBERLE  and  Vorländer,  19-^" 
HÄCKEL,  252 

Hahn  and  Trommsdorf,  E,.,  206 
Münchner  med.  Wochenschr,  No.  19 
(1900) 
Hailer,  E.,  199* 

Arbb.   d.  k.    Gesundheitssamts    29, 
H.  2  (1908) 
Hales,  66 
Halliburton,  219* 
Hamburg,  M.  and  Fraenkel,  S.,  190* 
Hamburger,  H.  J.,  236,  244,  254,  307, 
322,  322*2 
2)    Biochem.    Zeitschr.    11,    443-480 

(1908) 
and  Hekma,  287* 

Biochem.  Zeitschr.  3,  88-108;  7,  102- 
116  (1906-1907);  9,  275-306,  512- 
521  (1907);  24,  470-477  (1910) 
Koeppe,  H.  and  Overton,  E.,  236 
and  Limbeck,  von,  314 
and  Limbeck,  von  and  Gürber,  A., 
321 
Handovsky,  H.,  82*3,  ng^  147/«^  149*1^ 
151,  152*2,  155,  362*2  /« 

1)  Koll.-Zeitschr.  4  and  5  (1910) 

2)  Biochem.  Zeitschr.  25,  510  (1910) 
and  Pauh,  Wo.,   147^«,  149*i,    151, 

311 
Hannover,  421 

Harden,  A.  and  Young,  W.  J.,  191 
Proc.    Roy.    Soc.    77,    B,    405-420 
(1906);  78,  B,  369-375  (1906) 
Hardy,  W.  B.,  10,  64,  145,  158,  158*i, 
159,  159*2 
1)  Journ.  of  Phys.  33,  251-337 
Proc.  Roy.  Soc.  79,  413-426,  Ser.  B 
(London) 
Harlow,  M.  M.  and  Stieles,  P.  G., 
189* 
Journ.  of  Biol.  Chem.  6  (1909) 


Harnack,  E.,  142 

Harriman,  Mrs.  Oliver,  169-^'' 

Harris  and  Donnan,  46 

Harrison,  W.,  135* 

Harvey  Lectures,  210,  228^",  299 

Hatmaker  and  Just,  178 

Hatschek,  E.,  15, 15*,  16, 16*,  262,  348 

Koll.-Zeitschr.  6,  254-258  (1910);  7,. 
81-86  (1910) 
Hansen  and  Drey  er,  144 
Hausmann,  J.,  262* 

Zeitschr.  f.  anorgan.  Chemie  40,  110- 
145  (1904) 
Haversian,  263 
Hay,  413* 

Journ.  of  Anat.  and  Phys.  16  and  17 
Heald,  F.  D.,  382 
Hecker,  156,  157,  206 

and  Pauh,  156,  157 
Hedin,  S.  G.,  28*4,  185,  191*2,  192*,, 
243*1,  307 

1)  Pflüger's  Arch.  60,  360  (1895) 

2)  Biochem.  Journ.  1,  484-495  (1906)^ 

3)  Biochem.  Journ.  1,  474,  484;  2,, 
27,  81,  112  (1906) 

")  Zeitschr.  f .  physiol.  Chemie  50,  497' 
(1907);  60,  364  (1909) 
Heidenhain,  M.,  318 

Pflüger's  Arch.  56,  579  (1894) 
Heinz,  408 
Hekma,  160,  279 

and  Hamburger,  H.  J.,  287* 
Helbig,  435 
Held,  234,  234* 

Arch.  f.  exper.  Pathol,  u,  Pharm.  53;. 
227 
Helmholtz,  353 
Henderson,  310 

Lawrence   J.   and  Fischer,    M.   H.,. 
232^" 
Henri,  V.,  83*,  187 

Comptes  rend.  147,  62-65 

(and  Girard-Mangin),  204* 
Henry  (laws),  20,  242,  243,  308,  309, 

388,  396 
Herbst,  C,  264 

Mitterlungen  d.  zool.  Station  NeapeL 
11,  185,  191  (1803) 
Herman,  H.,  293 
Hermann  and  Dupre;,  Fr.,  348 

Pflüger's  Arch.  26,  442 


AUTHORS'   INDEX 


445 


Herzog,  B.  and  Adler,  27* 

R.  O.,  53,  53^",  54,  103,  104,  146, 

146*" 
Zeitschr.   f.   Elektrochcm.   13,    533- 

539  a  907) 
and  Arrhenius,  Sv.,  47 
(and  Graham,  Th.),  146*« 
and      Kasarnowski,       104*,      183  , 

190* 
Biochem.  Zeitschr.  11,  172  (1908) 
and  Öholen,  L.  W.,  53^" 
Hess,  W.,  114 

Münchner  med.  Wochenschr.,  No.  32 
(1907) 
Hessberg,  P.,  208,  209 
Heyden,  von,  99,  366,  376 
Hill,  A.  Croft,  250 
A.  C.  and  Parnas,  298 

HiROKOWA,  411 

Hersch  and  Beck,  113 
Hirschfeld,  M.  and  PauU,  152 
Hirshfeld,  L.,  204,  204*^,  285,  285*, 
286 

1)  Zeitschr.    f.    allg.    Phys.    9,    529- 
534 

2)  Arch.  f.  Hyg.  63,  237-286 
Höber,  R.,  81,  243*«,  244*^,  292*,  294*, 

295,  295*«,  295*",  296,  298,  307, 
307*^  311,  311*2,  319*1^  322,  329, 
353,  353*3",  382,  386,  387,  387** 

1)  Pfluger's    Arch.    70,    624    (1898); 
75,  246  (1899) 

2)  Pfluger's  Arch.  81,  522  (1900) 

•■>)  Physikal.  Chem.  d.  Zelle  (Leipzig, 

1906) 
3")  Physikal.  Chem.  d.  Zelle   u.    d. 

Gewebe  (Leipzig,  1911) 
*)  Pfluger's  Arch.  120,  508  (1907) 
5)  Physikal.  Chem.  u.  Physiol,  in  A. 

V.    Koränyi    u.    P.    F.    Richters 

Physikal.   Chemie  u.   Medizin   1, 

389  (Leipzig,  1907) 
«)  Biochem.  Zeitschr.  14,  209  (1908) 
")  Biochem.  Zeitschr.  14,  209  (1908) 

Pfluger's  Arch.  134,  311  (1910) 

8)  Biochem.  Zeitschr.  17,  518  (1909) 

9)  Zeitschr.  f.  Physikal.  Chemie  70, 
134-145  (1910) 

10)  Zeitschr.  f.  aUg.  Physiol.  10,  173- 
189  (1910) 


HöBKR,  R.,  ")  Arch.  .^  d.  ges.  Physiol. 
134,  311-336  (1910) 
and  Chassin,  S.,  427* 
KoU.  Zeitschr.  3,  76  (1908) 
and  Joel,  A.,  386 
and  Ruhland,  242 
and  Waldenberg,  295* 
Pfluger's  Arch.  126,  331  (1909) 
HoFF,  J.  H.  van't,  43 
Hofmeister,  F.,  67*,  95,   147^",  151, 
166,  220,  276,  277,  318,  319,  337, 
343,  409 
Arch.   f.   e.xper.   Pathol,   u.   Pharm 

27,  395  (1890^;  28,  210  (1891) 
and  Ostwald,  Wo.,  337 
and  Pick,  166 
P.,  115 
HoGAN,    J.  J.   and   Fischer,  W.  H., 

220''' 
Holde,  D.,  11*,  64* 

Chemiker.     Ztg.,    No.     54     (1908); 
Zeitsciir.  :'.  angew.  Chemie,  2138ff, 
(1908) 
Holderer,  187 
Holer,  R.,  230 
hollinger,  235* 

Biochem.  Zeitschr.  17,  1  (1908) 
Hooker,  D.  R.,  231,  332 

Am.    Journ.   of   Physiol.   27,    24-44 

(1910) 
Marian  O.  and  Fischer,  M.  H.,  33, 
36,  36^",  352 
Hopkins,  F.  G.  and  Fletcher,  298 
Hoppe  and  Seyler,  226 
Howell,  299 

HowLAND  and  Marriot,  314 
Hunt,  R.,  366 

Illyes,  G.  and  Kövesi,  340 

Inada,     R.     and     Müller,     O.,     311, 

380 
IscovESCO,  H.,  185*^  239,  299,  300*^, 

330*2,  342*2^  375/" 

1)  Comptes  rend,  de  la  Soc.  de  Biol. 
60  (1906) 

2)  Etude  sur  les  humeurs  de  I'organ- 
ique  (Paris,  1906) 

3)  Biochem.     Zeitschr.     24,      53-78 
(1910) 

Ishizaka  and  Schucht,  84*^ 
Isuneoka  and  Friedberger,  365 


446 


AUTHORS'   INDEX 


IzAR,  G.,  366*^  372*1,  375*2^  375^  370*2 

1)  Biochem.   Zeitschr.   20,    249,   266 
(1909) 

2)  Zeitschr.    f.    Immunitätsforschun- 
gen, Oris?.  2 

3)  Zeitschr.  f.  klin.  Medezin  68,  516 
and  Ascoli,  M.,  211,  365%  366,  371, 

373*,  377*,  383^" 

Jacobson,  P.,  187 
Jacoby,  M.,  184*,  196 

Biochem.  Zeitschr.  4,  21  (1907) 

and  Schütze,  A.,  189*,  196 

Zeitschr.   f.   Immunitätsforschungen 
4,  730-739  (1910) 
Jacque,  L.  and  Zunz,  E.,  199* 

Arch,  intern  de  Physiol.  8,  227-270 
(1909) 
Jagic,  N.  and  Landsteiner,  K.,  200*, 

204* 
Japelli,  G.  and  D'Errico,  296 
Jensen,  P.  and  Fischer,  H.  W.,  212, 
222* 

and  Fischer,  M.  H.,  291* 
Joel  and  Höber,  R.,  386 
Johnson,  184 
Joos,  J.,  201* 

Zeitschr.  f .  Hygiene  40,  203  (1902) 
JoRDis,  E.,  93,  93^"- 
Joseph,  377 

Dermat.  Zentrabl.  (Sept.  1907) 
JOSLIN,  314 
JosT,  L.,  259* 

Tharandter  forst.  Jahrbuch  60,  331- 
334  (1909) 

JUNGFLBISCH,  20 

JÜNGSEN  and  (Sörensen),  143* 
Just  and  Hatmaker,  175 

Kahlenberg  and  True,  382 
Kasarnowski,  H.  and  Herzog,  R.  O., 

104*,  183*,  190* 
Katz,  J.  R.,  178,  292 
Katzenellenbogen,  319* 

Pflüger's  Arch.  114,  522  (1906) 
Kaufmann,  M.,  366 
Keesom,  119 

KiLNiTZ  and  Gerloff,  263*''* 
Kirschbaum,  99,  102,  197,  205 

and  Pribram,  99 
Kisch,  240 


Klatte  and  Büchner,  184* 
Klemensiewicz,  231 
Klemperer,  G.,  343,  343-^" 
Klose  and  Vogt,  352 
Kneipp,  Fr.,  364 
Knoevenagel,  E.,  70,  187,  428 

Sitzung  d.  ehem.  ges.  zu  Heidelberii' 

17,   2    (1911);     (Zeitsclir.  f.  ang. 

Chan.  505  (1911) 
KoBER,  P.  A.,  120,  120^"- 
Kobler,  173 

Koch,  Robert,  138,  193,  395,  396 
Koeppe,  H.,  307 

and  Hamburger  and  Overton,  236 
Köhler,    F.    and    Traube,    J.,    163, 

427/» 

KOHLER,  R.,   95 

Kolle,  von,  196-'^" 
KoPACZEWSKi,  94,  94*"- 
KoRANYi,  A.  VON,  315,  339*,  341 

Zeitschr.  f.  Idin.  Medezin  34  (1898) 

BerUner  Idin.  Wochenschr.  781(1899) 

and  Bence,  J.,  315 

and  Kövesi,  341 

Kövesi  and  Roth-Schultz,  340* 

Pathol,    u.    Therapie    d.    Merenin- 
suffizienz  76  (Leipzig,  1904) 

and  Richter,  O.,  315 

Physikal.  Chemie  u.  Medizin  (Leip- 
zig, 1907-1908) 
K0RÄNYI,  P.  T.  and  Richter,  A.  v., 

113 
Kossel,  a.,  160,  279 

Münchner  med.  Wochenschr.  58,  2 
(1911) 
Kövesi  and  Illyes,  G.,  340 

A.  V.  and  Koränyi,  A.  von  and  Roth- 
Schultz,  340* 

andSuranyi,  341 
Krafft,  f.,  46 

and  Smits,  A.,  45,  46 
Krans,  313 
Kreidl,  a.  and  Lenk,  350* 

and  Neumann,  349* 
Kröenig,  50 

Krönig  and  Paul,  395,  401,  402,  403 
Kubowitsch,  J.  A.,  266 
Kunde,  216 
Kundt,  76 

Kupfer,  von,  248,  373 
KtJsTER,  E.,  264 


AUTHORS'   INDEX 


447 


Laer,  W.  H.  van,  180,  184 

T.  Congres  intern,  de  Brasserie;  25,  7 

(1910) 
Bull.  Acad.  R.  Belg.  305-320;  302- 
370  (1911) 
Lagergreen,  337 

Lamy,  E.  and  Mayer,  A.,  334*,  335 
Comptes  rend,  de  la  Soc.  de  Biol. 
222  (1904) 
Landstetner,    K.,    196^",    198,    200, 
201,  205 
and  Botteri,  A.,  199* 
Zentralbl.  f.Bakt.  42,  562-5G6  (190G) 
Eisenberg  and  Volk,  201 
and  Jagic,  N.,  200*,  204* 
Münchner  med.  Wochenschr.,  No.  27 

(1904) 
and  Pauli,  Wo.,  205*,  205 
Wiener   med.  Wochenschr.,   No.   18 

(1908) 
and  Uhlirz,  145* 

Centralbl.  f.  Bakter.  40,  206  (1905) 
and  Weleck,  St.,  200 
Zeitschr.  f.  Immunitätsfor.  u.  exper. 
Therapie  8,  397-403  (1910) 
XiANDWEHR  and  Baisch,  343 
Langley,  J.  N.  and  Fletcher,  328 
Laqueur,  E.,  154,  164 

and  Sackur,  O.,  154*,  164,  164* 
Beitr.  z.  chem.  Physiol  u.  Pathol.  3, 
193,  224 
Lawes  and  Gilbert,  265 
Lea,  Carey,  366 

Am.  Journ.  of  Science  (3),  37,  476; 
38,  47;  41,  482 
Leavenworth  and  Mendel,  266 
Lebedew,  a.  von,  102*,  190* 

Biochem.  Zeitschr.  20, 114-125  (1909) 
Leber,  Th.,  286 
Leclef  and  Denys,  288 
Leduc,    Stephane,    254,    254^",    255, 

255^"-,  256,  256^^'-,  257 
Lemanissier,  J.,  73*,  144,  144*,  349* 
L'Etude    des    corps     ultramicrosc. 

Thesis  (Paris,  1905) 
L'Etude     des     corps     ultramicrosc, 

Rousset  (Paris,  1905) 
and  Michaelis,  144 
Lenk,  E.,  169,  350* 
and  Fürth,  O.  von,  291 
andKreicU,A.,350* 


Lennep,  Ross  van,  181 
Lepeschikin,  W.  W.,  240,  245,  278 
Levaditi  and  Yamanouchi,  208 
Levites,  S.  J.,  162 
Lewith,  151 

Lichtwitz,  L.,  342*^,  343,  363,  364 
1)  D.    med.    Wochenschr.,    No.    15 

(1910) 
-)  Zeitschr.    f.    physiol.    Chemie   64, 

144-157  (1910) 
and  Rosenbach,  F.  J.,  342*,  343* 
Zeitschr.  f.  physiol.  Chemie  61,  117 
(1909) 
Lieber  and  Schmidt,  P.,  189 
Liebermann,  L.,  152*,  153*,  177 

L.  von,  206,  265 
Liesegang,    R.    E.,    106*i,    239,    257, 
260*3,    261,    261*1,    261'"-,     263, 
263*%  264,  269*,  269*^ 

1)  Chem.    Reaktionen    in    Gallerten 
(Leipzig,  1898) 

2)  Beitr.    z.    einer   koUoidchem.    d. 
Lebens  (Dresden,  1909) 

3)  Ann.  d.  Phys.  19,  406  (1906);  32, 
1095  (1910) 

4)  Koll.  Zeitschr.  7,  219  (1910) 

5)  Naturw.    Wochenschr.,     No.    41 
(1910) 

»)  Journ.  f.  Phychol.  u.  Neurol.  17, 
1-18  (1910) 
LiLLiE,  K.  G.,  378 

Am.  Journ.  of  Physiol.  17,  89  (1906); 

24,  459  (1909) 
R.  S.,  295*2,  354 

Am.     Journ.    of    Physiol.    10,    419 
(1904) 
Limbeck,  V.,  Gürber,  A.  and  Ham- 
burger, H.  J.,  321 
von  and  Hamburger,  314 
LiPMAN,  C.  B.,  379 

Loeb,  Jacques,  225^",  230,  241,  245, 
245*^  264*3,  282,  289*,  378,  379, 
413*2 

1)  Pflüger's  Arch.  69,   1  (1898);  71, 
457  (1898);  75,  303  (1899) 

2)  Pflüger's  Arch.  91,  248^(1902) 

3)  Untersuch,  üb.  d.  künst.  Parthe- 
nogenese (Leipzig,  1906) 

«)  Biochem.  Zeitschr.  11,  144  (1908) 
J.  and  Bentner,  R.,  240,  295,  295^" 
and  Osterhout,  W.  J.  V.,  241 


448 


AUTHORS'   INDEX 


LÖPPLER,  30 

LoEPER,  321*,  412* 

Comptes  rend,  de  la  Soc.  de  Biol. 
58,  1056  (1905) 
LoEWE,  G.,  243 

S.,  139,  386 
LoEWY,  A.,  308*2,  309,  312*i 

1)  in  A.  von  Koränyi  u.  P.  F.  Rich- 
ters Pathol,  d.  Respiration  2,  37 
(Leipzig,  1908) 

2)  in  A.  von  Koränyi  u.  P.  F.  Rich- 
ters Physikal.  Chemie  u.  Medezin 
1,  248  (Leipzig,  1908) 

and  Munzer,  E.,  313* 

Arch.  f.  Physiol.  (1901) 
LÖFFLER,  30,  30-^" 
Lorenz,  R.,  11,  49^",  51 
LosEv,      G.      and      Freundlich,      H., 

27* 
Lottermoser,  A.,  28*,  84*,  -365^" 

Koll.  Zeitschr.  6,  78-83  (1910) 
Lowe,  Chas.,  252^" 
Löwe,  H.,  362 
lubarsch,  231 
LuDERKiNG,  Chas.  and  Wiedermann, 

E.,  134*,  137* 
Ludwig,  C,  66,  336,  337 
Ludwig,  G.,  33r" 
Luther  (-Ostwald),  108^",  113 

Macallum,  a.  B.,  26,  234,  234-^",  287, 
292,  296,  296*,  298 
Science  (Oct.  7,  14,  1910) 
MacCallum,  413,  413* 

1)  Pflüger's  Arch.  104;  Am.  Journ.  of 
Physiol.  10,  101,  259 

2)  On  the  mechanism  of  the  physiol. 
action  of  the  cathartics  (Univ.  of 
Cal.  pubUc,  1906) 

MacDonald,  298A,  354 
McClendon,  J.  M.,  98,  379,  405 

MCCOLLUM,  351 

Madsbn,  Th.,  56,  105 
Magnus,  R.,  190,  190*,  227,  333*i 
1)  Arch.  f.    exper.    Pathol.   45,    210 

(1901) 
^)  Zeitschr.   f.   physiol.    Chemie  41, 
149-154  (1904) 
Magnus,   R.  and   Gottlieb,   R.,   332, 
336* 


Mai,  J.  and  Rothenpusser,  S.,  174 

1)  Milchwirfcschaftl.  Zeutralbl.,  No.  4 
(1910) 

2)  Zeitschr.  f.  Unters,  d.  Nahrungs- 
genussmittel, No.  12  (1909) 

Maier  and  Forges,  O.,  208 
Malfitano,  G.,  91,  93^'^-,  95,  102 

and  Duclaux,  J.,  92,  102 

and  Moschkoff,  A.  N.,  134* 

Comptes  rend,  de  I'acad.  d.  sciences 
151,  817ff. 
Mangin     and     Henri     and    Girard, 

204 
Mansfeld,  G.,  388* 

Pflüger's  Arch.  131,  457-464  (1910) 

and  Bosanyi,  386 
Marc,  120,  121,  180 

Rob,  26* 
Marchand,  23 

Margadant  and  Frei,  396,  403 
Marinesco,  266 
Marriot  and  Howland,  314 
Martin,  C.  J.,  143*,  146* 
Masius,  M.  and  Michaelis,  L.,  337 
Massard  and  Bordet,  J.,  286 
Masuda,  353 
Mathews,  353,  353*,  382,  382* 

Science  15,  492  (1902) 
Matruchot,  p.  and  Molliard,  216* 

Rev.  Zen.  de  Botanique  14  (1902) 
Matuta,  j.,  152,  156-^" 
Mauabe,  152 
Mayer,  Andre,  159,  159* 

Comptes  rend.  (Oct.  8,  1906) 

and  Edinger,  421 

A.  and  Lamy,  E.,  334*,  335 

and  Schaeffer,  G.,  279* 

Comptes  rend,  de  la  Soc.  de  Biol.  64, 
681  (1908) 
Mayerhofer,   E.   and  Pribram,   E., 
322* 

1)  Wiener  khn.Wochenschr.  25  (1909) 

2)  Zeitschr.  f.  exper.  Pathol,  u.  Thera- 
pie 7  (1909) 

3)  Biochem.  Zeitschrift  24,  453-469 
(1910);  27,  376-384  (1910) 

Mechowski,  W.,  363 
Mecklenburg,  W.,  25,  25*,  119,  120 
Meigs,  E.  B.,  290,  290*,  291* 

Am.  Journ.  of  Physiol.  26,  191-211 
(1910) 


AUTHORS'   INDEX 


449 


Meijeringh,  W.,  175* 

Chem.  Weekblivl  7,  951-953 
Meltzer,  S.  J.,  386,  387 

and  Shaklee,  A.  O.,  189* 
Mendel,  L.  B.  and  Leavenworth, 

266 
Merck,  99 
Merino,  von,  323 
Mbstrezat,  354 

Metcalf,  W.  v.,  33*,  34,  35,  347* 
Zeitschr.  f.  physikal.  Chemie  52,  1 

(1905) 
(and  Ramsden),  348 
Metchnikoff,  Elie,  285 

E.,  193,  239 
Meurer,  R.,  245* 

Jahrb.  f.  wessensch.  Botanik  46,  503- 

567  (1909) 

Meyer,  H.  and  Gottlieb,  R  ,  332*, 

380*,  412* 

Exper.  Pharmakologie  (Berlin,  1910) 

Hans  and  Overton,  E.,  385,  386,  387, 

388 
Kurt,  54* 

Hofmeisters  Beitr.  z.  chem.  Physiol, 
u.  Pathol.  7,  393Ef. 
Michaelis,  L.,  27,  28,  77,  107,  113, 
119,  119'"-,  141,  144,  144*1,  145^ 
147,  147*3,  i5§^  161^  164^  165^ 
185*2,  186,  187,  201,  202,  285 

1)  D.  med.  Wochenschr.  42  (1904); 
Virchows  Arch.  179,  195-208 
(1905) 

2)  Biochem.  Zeitschr.  7,  488-492;  12, 
26;  16,  81-86,  486-488;  17,  231- 
234;  19,  181-185  (1907-1909) 

3)  Biochem.  Zeitsclu".  19,  181  (1909) 
and  Beniasch,  204 

and  Davidsohn,  202 
and  FremidUch,  79 
and  Lemanissier,  J.,  144 
and  Masius,  337 

andRona,  P.,  107,  107*^  141*i,  145*, 
147,  201,  234,  348* 

1)  Biochem.  Zeitschi".  4,  11  (1907) 

2)  Biochem.  Zeitschr.  14,  476ff  (1908) 
»)  Biochem.  Zeitschr.  15,  196  (1908) 
■*)  Biochem.  Zeitschr.  21,  114-122 

5)    Biochem.   Zeitschr.   25,    259-^36() 

(1910) 
and  Skwirsky,  206 


MicuLiciCH,  M.,  307* 

Zcntralbl.  f.  Physiol.  24,  12 

Minkowski,  O.,  314 

MiTTELBACH,  Robert,  92-'^" 

Modelski,  J.  W.  and  Pfeiffer,  156 

MoLiscH,  H.,  216,  217^"- 

Moll,  159* 

Hofmeisters  Beitr.  4,  563  (1903) 

MoLLiARD  and  Matruchot,  P.,  216* 

Moore,  A.  R.,  230 
and  Roaf,  H.  E..  387* 
B.  G.  and  Roaf,  H.  E.,  73* 
Proc.  Roy.  Soc.  of  London,  Ser.  B. 
73,  382  (1904);  77,  86  (1906) 

MoRAWiTz,  P.,  394* 
Kolloidchem.  Beihefte  1,  301  (1910) 

Morgan,  J.  L.  R  ,  109 

Morgenroth,  J.,  201*i,  205*^ 

1)  Münchner  med.  Wochenschr.,  No. 
2  (1903) 

2)  Arbb.  a.  d.  Pathol.  Inst.  (Festschr.) 
(Berlin,  1906) 

and  Pane,  D.,  206*^,  206* 
Biochem.  Zeitschr.  1,  354-366  (1906) 
Mörner,  K.  a.  H.,  72*,  343*2 

1)  Zeitschr.  f.  phj'-siol.   Chemie  34, 
207  (1901) 

2)  Skandinav.  Arch,  f .  Physiol.  (1901) 
MoROCHOWETz,  Leo,  94,  94*"- 

Morse,  H.  W.  and  Pierce,  G.  W., 

262* 
Zeitschr.    f.    physikal.    Chemie    45, 
589-607  (1903) 
MoRUZzi,  G.,  311* 

Biochem.  Zeitschr.  28,  97-105  (1910) 
MoscHKOFF,    A.    N.    and    Malfitano, 

134* 
Motzfeld,  339* 
MOUFANG,  E.,  180 
Much,  H.,  144,  196 

Römer  and  Siebert,  C,  144* 
Zeitsclir.  f .  cüätet.  u.  physikal.  Thera- 
pie 8  (1904,  1905) 
Mühlmann,  266 

MÜLLER,  O.  and  Inada,  R.,  311,  380* 
Deutsch,  med.  Wochenschi-.  (1904) 
and  Thürgan,  H.,  216,  256*,  257^"' 
Zentralbl.    f.    Bakter.    20    (II)    No. 
12/14;  51/17  (1908) 
Munch,  296 


450 


AUTHORS'   INDEX 


MuNK,  F.,  208 

M.,  264 
MÜNTZ,  A.,  215* 

Comptes  rend,  de  I'acad.  d.  sciences 
150,  1390-1395  (1910);  151,  790- 
793  (1910) 
MuNZER,  E.  and  Loewy,  A.,  313* 

MUTH,  W.,  112* 

Nagel,  G.,  33* 

Ann.  d.  Physik.   (4)  29,   1029-1056 
(1909) 
Nägeli,  a.  E.,  215,  216 

O.,  278 
Nailashima,  247 
Nathan,  E.  and  Sachs,  H.,  210 
Nathanson,  240 
Neisser,  M.,  86,  202*,  435 

Hys;ien.      Rundschau      13,       1261 

(1Ö03) 
and  Friedemann,  U.,  84*,  86,  203, 

203*,  205*,  283* 
Mi'mchner   med.  Wochenschr.,   No. 

11,  19  (1904) 
and  Guerrini,  288* 
Arbb.  a.  d.  Kgl.  Inst.  f.  exper.  Thera- 
pie, No.  4  (1908) 
and  Sachs,  A.,  207 
N'ell,  Peter,  54* 

Drude's  Ann.  18,  323  (1905) 
Nernst,  W.,  20,  63,  122 
Neubauer,  E.  (and  Porges,  O.),  87*, 
140*,  141*,  388* 
O.,  208 
Neuberg,  C.,  381,  381* 

1)  Sitzg.  d.  D.  Chem.  Ges.  v.  VI,  7 
(1904) 

2)  Biochem.  Zeitschr.  1,  166  (1906) 

3)  Koll.  Zeitschr.  2,  321,  354  (1908) 
and  Albu,  A.,  218*,  219*,  233* 

Neufeld,  288 
Neumann,  A.,  247* 

Zeutralbl.  f.  Physiol.  21,  102-105 

and  Kreidl,  A.,  349* 
NiCLOux,  M.,  388* 

Les  anesthesiques  generaux   (Paris, 
1908) 
NoFF,  299 
NovY,  J.,  328 

and  Vaughn,  210 


Obermeyer,  Fr.  and  Pick,  E.  P.,  197* 

Wienerkhn.  Wochenschr.  (1906) 
O'Connor  and  Gros,  O.,  365*,  372* 
Oden,  S.,  80 

and  Ohion,  Sl^"" 

and  Pauli,  Wo.,  152 
Oesper,  17^" 
Ohlon  and  Oden,  81-^" 
ÖHOLM,  L.  W.,  53,  53^",  54, 103*,  103*"" 

Zeitschr.  f .  physikal.  Chemie  50,  309- 
349  (1904) ;  70,  278-407  (1909.^ 

and  Herzog,  R.  O.,  53^" 
Qkbr-Blom,  M.,  307,  319* 

Skandinav.  Arch.  f.  Physiol.  20,  102- 
114  (1907) 
Okolska  and  Eisenberg,  403 
Omorvkow  and  Sachs,  H.  and  Ritz^ 

196 
Onnes,  Kamerlingh,  119 
Oppenheimer,  C,  182 

Kurt,  174 
OsTERHOUT,  W.  J.  v.,  241,  241^«,  379,. 
386 

1)  Botanical  Gazette  46,  53-55  (1908) 

2)  Journ.  of  Biol.  Chem.  I,  363-369- 
(1906) 

and  Loeb,  J.,  241 
OsTWALD,  Walter  and  Riedel,  A.,  179 
Wilhelm,  5^",  28*,  64,  113,  178,  259, 
262*2 

1)  Zeitschr.  f.  physikal.  Chemie,  62^ 
512  (1908) 

2)  Lehrt,  d.  allgem.  Chemie  2,  11 
(2"<^  ed.) 

Wolfgang,  12,  13,  17^",  26,  66,  67*\ 
70,  71^'^,  87,  147^",  161,  309,  309*3, 
310,  379,  379*1,  330,  sgO** 

1)  Pflüger's  Arch.  106,  568  (1905) 

2)  Pflüger's  Arch.  108,  563  (1905) 

3)  Koll.  Zeitschr.  2,  264,  294  (1908) 

4)  Koll.  Zeitschr.  6,  297  (1910) 
and  Freundlich,  147-^" 

and  Hofmeister,  337 

Luther,  108^'^,  113 

Sprengel,  113 
Oswald,  A.,  232*,  233 

Zeitschr.  f .  exper_Pathol.  u.  Therapie 
8,  2ß6  (1910) 
Ottenberg,  R.,  190 
Ottolenghi,  408 

Desinfektion  2,  109 


AUTHORS'   INDEX 


451 


Overton,  C.  E.,  66*,  236,  239,  240,  241, 
242,  289*,  294,  294*,  353 

1)  Pflüger's  Arch.  92,  115  (1902) 

2)  Pfliiger's  Arch.  105,  176  (1904) 

3)  Biochem.  Zeutralbl.  2,  518 

«)  Verh.  d.  Ges.  D.  Naturf.  II,  416 

(1903) 
and  Hamburger  and  Koeppe,  236 
and   Meyer,    Hans,    385,    386,    387, 

388 

Paal,  C,  31,  86,  99 

Padtberg,  234 

Pane,   D.  and  Morgenroth,   J.,   206*, 

206*5 
Paneth,  88 
Park,  359 

Parnos  and  Hill,  A.  V.,  298 
Parodko,  Th.,  282 
Pasteur,  L.,  193 
Patin,  G.  and  Roblin,  L.,  373* 

Journ.    Pharm,   et   Chemie    (6)    30, 
•    481-483 
Paul,  H.,  222* 

Mittlgn.  d.  k.  Bayr.  Moorkulturan- 

stalt  No.  2  (1908) 
Th.,  366 

and  Krönig,  395,  401,  402,  403 
Pauii,  Wo.,  67*S  68,  82,  86,  113,  115, 
137*S  147,  147^",  148,  149*S  151, 
152,  154,  156,  156*S  157,  205,  293, 
343,  381**,  410*3,  413*3 

1)  (Pascheles)    Pflüger's    Arch.    67, 
225  (1897) 

2)  Pflüger's  Arch.  67,  219  (1897);  71, 
1  (1898) 

3)  Verh.  d.  Kong.  f.  inn.  Med.  21, 
396  (1904) 

Sitzungber.  d.  k.  Akad.  d.  Wiss.  113 
(1904);  115  (1906) 

4)  Verh.  d.  21  Kong.  f.  inn.  Med. 

5)  KoU.  Zeitsclir.,  No.  5,  1,  241  (1910) 
and  Handovsky,   H.,    147^",    149*i, 

151,  311 

1)  Beitr.  z.  chem.  Physiol,  u.  Pathol. 
9,  419 

2)  Biochem.  Zeitschr.  18,  340ff .  (1909) 

3)  Biochem.    Zeitschr.    24,    239-262 
(1910) 

and  Hecker,  156,  157 
and  Hirschfeld,  M.,  152 


Pauli  and  Landsteiner,  205,  205* 

and  Oden,  S.,  152 

and  Rona,  162,  162* 

Hofmeisters  Beitr.  z.  chem.  Physiol, 
u.  Pathol.  2,  Iff. 

andSamec,  148*,  154,  161*,  268,  268* 

Biochem.      Zeitschr.      17,      235-256 
(1909) 
Pekelharning,  C.  a.,  286 
Pelet  and  Jolivet,  L.,  427* 

Die     Theorie     d.     Färbesprozesses 
(Dresden,  1910) 
Pemsel  (and  Spiro),  152* 
Perkin,  Sir  Wm.  Henry,  252-^" 
Perrin,  J.,  9,  51,  52"'-,  78,  79 
Pettenkofer,  352 
Pettibone  and  Abderhalden,  185* 
Pfaundler,  M.,  268 
Pfeffer,  W.,  57*i,  221*=,  221^",  238*2, 
286 

1)  Osmot.  Untersuchungen  (Leipzig, 
1888) 

2)  Pflauzenphysiol.  (Leipzig,  1897) 
Pfeffer,  W.  and  Vries,  H.  dE,  239 
Pfeiffer,  P.  and  Modelski,  J.  W.,  156 

and  Wittka,  156 
Pflüger,  341 
Philippi,  E.,  365* 
Philippson  and  Demoor,  J.,  298* 
Pick,  A.,  166,  167^",  362^" 

and  Auerbach,  330 

and  Hofmeister,  166 

E.  P.  and  Obermayer,  Fr.,  197* 
•  Pickering  and  Plateau  and  Quincke, 
36-''" 
Pierce,  G.  W.  and  Morse,  H.  W.,  262* 
PiNcussoHN,  L.,  144*,  230,  384 

Biochem.  Zeitschr.  10,  356  (1908) 
Plateau,    Quincke    and   Pickering, 

36-^"  and  Poggendorff,  33 
Poggendorff  and  Plateau,  33 
PoNFiCK,  E.,  333 
ponomarew,  279 

Porges,  O.,  87,  140,  141,  196^",  203, 
208 

and  Maier,  208 

and  Neubauer,  E.,  87*,  140*,  141*, 
388* 

1)  Biochem.    Zeitschi-.    7,    152-177 
(1907) 

2)  Koll.  Zeitsclir.  5,  4  (1909) 


452 


AUTHORS'   INDEX 


Portig,  P.,  365* 

Dissertation  (Leipzig,  1909) 
POSNYAK,  E.,  116,  175 
POTTEVIN,  250 

Prescott,  Samuel,  182 
Preti,  365* 

Comptes  rend.  d.  la  Soc.  de  Biol.  65, 

52,  224;  Biochem.  Zeitschr.  (1909); 

Zeitschr.   f.   physiol.    Chemie   58, 

539;  60,  317 
Pribram,  B.  E.,  99,  102,  191*,  219*i, 

221*1,  222*2,  296,  327*^ 

1)  KoUoidchem.  Beihefte  2,  1  (1910) 

2)  Wiener    klin.     Wochenschr.     15 
(1911) 

and  Goldschmidt,  387* 
and  Kirschbaum,  99 
and  Mayerhofer,  322* 
and  Stein,  E.,  192 
Pringsheim,  H.,  57*,  105*,  135*,  260 
Jahrb.  f.  w.  Botanik,  28,  1-38  (1895) 
and  Eissler,  135 
and  Stoffel,  105* 

Quagliariello,  G.,  412* 

Biochem.  Zeitschr.  27,  516-530 
(1910) 
Quincke,  G.,  10,  18,  25,  35*2,  sg/»^ 
76*3,  78^  108^»,  137,  161,  300,  347, 
347*2,  354 
^)  Poggendorff's  Annalen  139,  Iff. 
(1870) 

2)  Wiedem.  Ann.  35,  590 

3)  Ann.  d.  Phys.  1  (Appendix  IV), 
85-86  (1902) 

and  Plateau  and  Pickering,  36-''" 

Rabl,  H.,  264 

Naturforschervers.     München     (cit. 

by  Liesegang)  (1899) 
RÄHLMANN,  E.,  73*1,  127^  136*1,  144, 

144*2 

1)  Berhner  kUn.  Wochensch.,  No.  8 
(1904) 

2)  Münchner  med.   Wochensch.  48 
(1903) 

3)  Arch.  f.  d.  ges.  Physiol.  112,  128- 
171  (1906) 

Rakowski,  a.,  133* 
Ramsay,  Sir  Wm.,  54 


Ramsden,  W.,  34*,  347* 

Arch.  f.  Anat.  u.  Physiol.  (Physiol. 

Abt.)  517  (1894) 
Zeitschr.  f.  physikal.  Chemie  47,  336 

(1904) 
and  Metcalf,  348 
Ranvier,  287 
Reichardt,  230,  231 
Reichel,  395* 

Biochem.    Zeitschr.    22,     149,    177, 
201  (1909) 
Reichenbach,  H.,  404,  408,  408-''" 
Reicher,  K.,  247* 

Deut.  med.  Wochensch.  34  (2),  1529 
(1908) 
Reid,  E.  W.,  47 
Reinders,  E.,  36,  238 

Kon.    Akad.    van     Wetenschappen 

Amsterdam.  Proc.  563-573  (1910) 

Reinhold,  B.  and  Riesenfeld,  E.  H., 

80 
Reinke,  J.,  66*,  116,  116*,  116'""- 

Hansteinsbotan.  Abhand.  4,  1  (1879) 
Rettger,  299 

Revello-Alves  and  Benedicenti,  157 
Rhumbler,    L.,    283-^",    284,    285 
Richter,  A.  von  and  Koranyi,  P.  T., 
113 
O.  and  Koranyi,  315 
Richter,  B.  F.  and  Roth,  W.,  341 
Riecke,  E.,  42 

Riedel,  A.,  and  Ostwald,  Walter,  179 
Riesenfeld,  E.  H.  and  Reinhold,  B., 
80* 
Zeitschr.    f.    physikal.    Chemie    66, 
672-686  (1909) 

RiESMAN,  228 

Ringer,  W.  E.,  152^'^ 

and  Adler,  379 
Ringers,  137 
RiTz,  189,  196 

and  Sachs,  H.  and  Omarvkow,  196 
RoAF,  H.  E.  and  Moore,  A,  R.,  387* 

and  Moore,  B.  G.,  73* 
Robertson,  T.  B.,  64*,  164,  164*,  166, 
377 

1)  Journ.  of  Physic.  Chem.  11,  542 
(1907);  12,  473  (1908) 

2)  Koll.  Zeitschr.  7,  7-10  (1910) 

3)  Die  Physikal.  Chemie  d.  Proteine 
(Dresden,  1911) 


AUTHORS'  INDEX 


453 


Robin,  A.  and  Weill,  E.,  371 
RoBLiN,  L.  and  Patin,  G.,  373* 
RoDEWALD,  H.,  6G,  134,  134*1 

Zeitschr.  f .  physikal.  Chemie  24,  193 
(1897) 
RoDOLico  and  Filippi,  372 
Roentgen,  18 
ROGEE,  220^'^ 
Rogers,  Allan,  169-''" 
Rogoff,  J.  M.,  300 
RoHDE,  E.,  33* 

Ann.  d.  Phys.  (4)  19,  935  (1906) 
RoHLOFF  and  Schinja,  162 
ROHONGI,  184 

romanowsky,  434 
Romberg,  E  ,  380 

Römer  and  Siebert,  C.  and  Much  144* 
RoNA,  P.,  107*2,  108,  115*,  141*1,  145*^ 
147,  201 

and  Michaelis,  L  ,  348* 

and    Michaelis,    107,    107*^,    141*i, 
145*,  147,  201,  234 

and  PauH,  162,  162* 

Biochem.  Zeitschr.  21, 114-122  (1909) 

and  Takahashi,  D.,  235* 

1)  Biochem.     Zeitschr.     30,     99-109 
(1910) 

2)  Biochem.    Zeitschr.    31,    33G-344 
(1911) 

RoNDONi,  P.,  206,  206* 

Zeitschr.  f.  Immun,  u.  expcr.  The- 
rapie 7,  515-543  (1910) 
and  Sachs,  H.,  208 
Röntgen,  W.  K.,  81,  144 
RoozEBOM,  H.  W.  Bakhuis,  6 
Rosenbach,  F.  J.  and  Lichtwitz,  L., 
342*,  343* 

ROSENTHALER,  L.,  183*,  188 

Biochem.  Zeitschr.  26,  9  (1910) 
ROSHARDT,  P.  A.,  238* 

Beitr.  z.  Botan.  Zentralbl.  25,  Abt.  I, 
243-357  (1910) 
Roth,  W.  and  Richter,  P.  F.,  341 
and  Strauss,  H.,  323* 
Zeitschr.  f.  khn.  Medizin  37,  No.  | 
RoTHE,  A.,  28 

Rothenfusser,  S.,  174, 174*,  175, 181* 
Zeitschr.  f.  Unters,  d.  Nahrungs  u. 
Genussm.  18,  135-155  (1909);  19, 
261-268,  465-475  (1910) 
and  Mai,  174* 


Rothmund,  V.,  64* 

RoTH-ScHULTZ      and     Koränyi     and 

Kovesi,  340* 
Roux,  W.,  110,  256-''" 
and  Yersin,  110  198* 
Ann.  de  I'lnst.  Pasteur  3,  273-288 
(1889) 
Rubner,  265 
RuHLAND,  241,  242,  427 

and  Höber,  242 
Rumpf,  219* 

Münchner  med.  Wochenschr.,  No.  9 
(1905) 
Runge,  E.  F.  (F.  E.),  252,  253 
RuNNSTRÖM,    J.    and    Backmann,    L., 

265,  265* 
Russell,  H.  L.  and  Babcock,  175* 
Rysselberghe,  B.  VAN,  243* 

Mcm.  de  l'Acad.  roy.  de  Belg.  58 
(1899) 

Sabbatani,  L.,  363,  376 
Sachs,  H.,  196,  198 

and  Altmann,  206,  208 

and  Neisser,  M.,  207 

Omorvkow  and  Ritz,  196 

and  Nathan,  E.,  210 

and  Rondoni,  P.,  208 

W.,  216 
Sackur,  164 

O.  and  Laqueur,  154*,  164,  164* 
Salkowski,  343 

Berliner  klin.  Wochenschr.,  No.  51, 
52  (1905) 
Sansum,  143,  383 
Santesson,  298* 

Skandinav.  Arch.  Phys.  14,  1  (1903) 
Salomon,  208 
Samec,  M.,  133,  135*,  148*,  154,  156 

and    PauU,    148*,    154,    161*,    268, 
268* 
Sasaki,  Kumoji,  342* 

Hofmeisters  Beiträge  9,  386  (1907) 
Savare,  M.,  342* 

Hofmeisters  Beiträge  9,  401;  11,  71 
Savarese  and  d'Errico,  330 
Schade,  H.,  231,  266,  343,  344* 

1)  Kell.  Zeitschr.  4,  175-261  (1909) 

2)  Kuli.  ehem.  Beihefte  1,  375  (1910) 
^)  Münchner  med.  Wochenschr.,  No. 

1   2  (1909) 


454 


AUTHORS'   INDEX 


Schade,  H.,  *)  Zeitschr.  f.  exper.  Path. 

■  u.  Therapi  8,  2-34 
ScHAEPFER,  G.  and  Mayer,  A.,  279 
SCHANZ,  144 
SCHEITLXN,  W.,  310*,  311 

Dissertation  (Zurich,  1909) 

SCHELLENS,  W.,  400 

Inaug. -Dissert.  (Strassburg,  1905) 
ScHEURLEN  and  Spiro,  395,  401,  402 
ScHiNYA  and  Rohloff,  162 
Schleicher  and  Schüll,  92,  94,  95-''", 

96 
Schleicher,  95,  96 
Schmidt,  C.  G.,  26* 

P.,  196,  205^" 

and  Lieber,  189 

Nielsen,  Signe  and  Sigval,  34*,  189* 
Schneider,  J.,  81 
Schoep,  a.,  95* 

Bull,  de  la  soc.  chem.  de  Belg.  24, 
10  (1910) 
Schoep,  A.,  92^"- 
Schönborn,  S.  329 
Schorr,  K.,  152,  154,  166 
Schroeder,  p.  von,  162,  361 
ScHUCHT  and  Freundlich,  25* 

and  Ishizaka,  84*^ 
ScHÜLL,  95,  96 

and  Schleicher,  92,  94,  95^",  96 
Schulz,  Fr.  N.  and  Zsigmondy,  E,.,  143 
ScHUMBERG  and  Zuntz,  N.,  216 
Schütz,  J.  and  Fürth,  H.  von,  191* 
Schütze,  H.  and  Jacoby,  M.,  196 
Schwarz,  C.,  291*,  294* 

Pfiüger's  Arch.  117,  161  (1907) 

SCHWENKENBECHER,  A.,   324*,  345 

Arch.  f.  Anat.  u.  Physiol.  121  (1904) 
Seddig,  50 
Seyler  and  Hoppe,  226 

SHAKLEE,A.O.andMELTZER,  S.J.,189* 

Am.  Journ.  of  Physiol.  25  (1909) 
Sherman,  H.  G.,  169^" 
Sholto,  J.  and  Drey  er,  28* 

and  Douglas  and  Dreyer,  200* 
SlEBECK,    R.,    336 
SlEBERT,  C.,  144*,  196 

and  Römer  and  Much,  144* 
SiEDENTOPF,    H.,     76*1,    122,     123"'- 
126^"-,  127 

KoU.  Zeitschr.  6,  No.  1  (1910) 

and  Zsigmondy,  R.,  6,  75 


Signe  and  Schmidt-Nielsen,  Sigval,. 

34*,  189* 
Sjöquist,  152* 

Skwirsky  and  Michaelis,  206 
Smoltjchowski,  M.  von,  50,  51 
Smits,  a.  and  Krafft,  F.,  45,  46 
sobieranski,  411 
Söhngen,  181 
Sellman,  Torald,  337 
Sörensen  and  Jürgsen,  143* 
Southard  and  Gay,  377 
Spiro,  K.,  67* 

Hofmeisters  Beitr.  z.  chem.  PhysioL 

5,  276  (1904) 
and  Bruno,  J.,  401*,  403* 
Arch.  f.  exper.  Pharmak.,  41 
and  Ellenger,  299 
and  Pemsel,  152* 
Zeitschr.  f.  physiol.  Chemie  26,  233. 

(1898) 
and  Scheurlen,  395,  401,  402 
Sprengel  (-Ostwald),  113 
Stahl,  E.,  287 

and  de  Bary,  286 
Starling,     E.     H.,     47,     332,     335,. 

336* 
Steche  and  Waentig,  184 
Stefan,  146 

Stephan  and  Graham,  54 
Stein,  E.  and  Pribram,  192 
Steiner,  H.,  28 
Stern  and  Battelli,  386 
Stiles,  P.  G.  and  Harlow,  189* 
Stodel,  G.,  365*,  371,  371^" 

Les  Coll.  en  Biol,  et  en  Ther.  These- 
(Paris,  1908) 
Stoeltzner,  W.  and  Dekhuysen,  C.,. 

420 
Stoffel,  F.,  55*,  74,  172*,  262 
Inaug.-Diss.  (Zürich,  1908) 
G.,  361*,  362* 
and  Pringsheim,  105* 
Stöhr,  291^"- 

Strassburg  and  Ewald,  R.,  226 
Strassburger,  E.,  238 
Straub,  W.,  360*,  361* 
Pfiüger's  Arch.  98,  5/6 
and  Freundlich,  243 
Strauch,    F.    W.    and    Abderhalden^ 
187 


AUTHORS'   INDEX 


455 


Strauss,  E.,  15ö 

H.,  323*1,  329,  329*=,  339,  341,  345 

1)  Zeitschr.  f.  klin.  Med.  57,  No.  f 

2)  in  Koränyi  and  Richter  2,  110 
and  Roth,  W.,  323* 

Streitmann  and  Fischer,  M.  H.,  296 

Stumpf,  364 

SvEDBERG,  Th.,  4,  5,  7,  9,  10,  48,  50, 

51,  53,  365^» 
Koll.  Zeitschr.  4,  169  (1909);   5,  318 

(1909) 
Sykes,  a.  and  Fischer,  M.  H.,  334 
Szücs,  J.,  382 

Tachau,  p.,  231,  232 

Taenzer  and  Unna,  P.  G.,  434 

Tait,  J.,  307^" 

Takahashi,  D.  and  Rona,  P.,  235* 

Tamara,  342* 

Arch.  f.  exper.  Pathol,  u.  Pharmek. 
59,  1  (1908) 

Tamman,  G.,  57* 

Wiedem.  Ann.  34,  299  (1888); 
Zeitschr.  f.  physikal.  Chemie  10, 
255ff.  (1892) 

Tangl,  f.,  18*,  266 

in  Oppenheimers  Handbuch  d.  Bio- 
chemie 3,  II,  p.  20 

Tappeiner,  363 

Teague,  O.  and  Buxton,  B.  H.,  84*^, 
430*2 

1)  Journ.ofexper.Med.9,No.3(1907) 

2)  Zeitschr.  f.  physikal.  Chemie  60, 
469-506  (1907);  62,  287-307  (1908) 

(and  Field),  205* 

Thomas,  A.  W.,  3 
Hay  ward,  G.,  228 

Thompson,  d'Arcy  W.,  281-''" 
and  Walti,  362 

Thouery,  363 

Tigerstedt,  R.  a.  a.,  293 

Translator,  3,  12,  26,  33,  40,  41,  42, 
48,  54,  71,  86,  98,  109,  120,  122, 
137,  143^",  145,  169,  175,  188,  190, 
195^",  199,  210,  210-^",  211^", 
220-^",  228,  230,  232,  232-^",  234, 
247,  252,  261,  276,  281-^",  287, 
298B,  299,  300,  308,  310,  323,  324, 
325,  335,  336,  338,  339,  350,  351, 
354,  359,  361,  365,  371,  377,  378, 
379,  383,  388,  404-405,  409,  435 


Traube,  J.,  108,  100, 163,  211,  236*'  ^'', 
236-^'^,  385*  ■^",  386 

Arch.  f.  Anat.  u.  Physiol.  87  (1867) 

Pflügers  Arch.   105,   541-558;   559- 
572  (1904) 

and  Czapek,  F.,  240,  386 

and  Kohler,  F.,  163,  427-'''' 

Moritz,  57* 
Trommsdorf,  R.  and  Hahn,  206 
Tröndle,  a.,  245 
True  and  Kahlenberg,  382 
TsuBoi,  298 
Tyndall,  119,  125 

Uhlenruth,  B.  D.,  257* 
Uhlirz  and  Landsteiner,  145* 
Unna,  P.  G.,  361*^  364*^ 

1)  Medizin,  Rev.  1,  No.  2-4 

2)  Medizin.  Rev.  Klinik,  No.  42,  43 
(1907) 

3)  Arch.  f.  mikrosk.  Anat.  (Waldeyer 
Festschr.)  78  (1911) 

and  Ehrlich,  424 

and  Golodetz,  L.,  163,  434 

and  Taenzer,  434 

Van  Slyke,  Donald,  210-^",  310,  314 
Van't  Hoff,  J.  H.,  43 
Vaughan  and  Novy,  210 
Vegesack,  H.  von,  73*,  106,  106* 
Vernon,  240 
Verworn,  388 
VoELTz,  346*,  347,  348 

Arch.  f.  d.  ges.  Physikol.  102,  373- 
414 
VoGT  and  Klose,  352 
Voigt,  J.,  366,  373,  374 
Voigtländer,  106 
Volk,  200,  201 

and  Eisenberg,  200,  200*,  203 

and     Landsteiner     and    Eisenberg, 
201 
Volkmann,  A.  W.,  218* 

Ber.  d.  Kgl.  sachs.  ges.  d.  Wissensch. 
(1874) 
Vorländer  and  Häberle,  19^" 
Vries,  H.  de  and  Pfeffer,  W.,  239 

Waentig  and  Steche,  184 
Wagner  and  Determeyer,  343* 
Walden,  p.,  57* 


456 


AUTHORS'   INDEX 


Waldenberg  and  Höber,  A.,  295* 
Wallace,  G.  B.  and  Gushing,  A.  R., 

319* 
Wallerstein,  180 
Walti  and  Thompson,  362 
Warburg  and  Wiesel,  386 
Wasielewski,  Th.  v.,  418 
Wasserman,  194,  196^",  206,  207,  208 
Webster,  R.  W.,  289 

Univ.   of   Chicago   Publ.   10    (Dec, 
1902) 
Weevers,  Th.,  234 
Wegelin,  G.,  4 
Weigert,  434 
Weil,  R.,  210 
Weill,  E.  and  Achard,  371 

and  Robin,  A..  371 
Weimarn,  p.  p.  von,  71,  73 
Weissmann,  375-''" 

Weleck,  St.  and  Landsteiner,  K.,  200 
Welter,  250 
WiDAL,  Fernand,  202^",  339 

and  Gruber,  202^" 
WiDMARK,  E.  M.,  222*,  292* 

Skandinav.  Arch.  f.  Physiol.  23,  421- 
430  (1910);  24,  13-22  (1910);  24, 
339-344  (1911) 
Wiedemann,  E.  and  Ludeking,  Chas., 
134*,  137* 

Wiedemann's  Annalen  25,  433 

G.,  78 
Wiegner,  G.,  346,  348,  349* 
Wiesel  and  Warburg,  386 

WiLKE,   119 

WiLKE-DöRFÜRT  and  Zsigmondy  and 

Galecki,  98 
Williams,  Mattieu,  169^" 
Winkel,  R.,  123 

WiNKELBEECH,  3S* 

Winter,  329 

WiSLiCENUs,    H.,    110,    112^"-,    169*, 
248*,  249,  250,  258*i,  258^^^- 

1)  Papier-Ztg.  16  (1910) 

2)  Tharandter  forst.  Jahrt.  60,  313- 
358 

3)  KoU.  Zeitschr.  6,  17,  87  (1910) 
and  Muth,  W.,  112* 
CoUegium  No.  255,  256  (1907) 


Wistinghausen,  63 
Witte,  35 

Wittka  and  Pfeiffer,  156 
WÖHLER,  L.,  73* 
Woodyatt,  R.  T.,  228,  228^" 

and  Fischer,  M.  H.,  335 
Wright,  288 


Yamanouchi  and  Lavaditi,  208 
Ybrsin  and  Roux,  110,  198* 
Young,  W.  J.  and  Harden,  A.,  191* 

Zangger,  H.,  55,  56,  63,  109,  172,  173, 
173*2,  196^" 

1)  Ergebnisse  d.  Physiol.  VII  (1908) 

2)  Schweizer.  Arch.  f.  Tierheilkunde 
5  (1908) 

Zeiss,  Carl,  120 

ZiEGLER,   J.,   10,   34,   73,   106,    115*2, 
138*2,  147^^*,  148*3 
and  Bechhold,  H.,  10*,  35,  54*2,  55^ 
55*2,  57*1^  73^   106,   138*2,  147^", 
148*3,  162*2,  238*1,  243,  244,  260*i, 
269*3,  319,  320*2,  327,  329,  343, 
378*3,  380,  403*2 
Kurt,  231 
ZiLLESSEN,  H.  and  Araki,  F.,  226 
Zlobicki,  135*,  137* 

BuU.  de  I'Acad.  de  Science  de  Cracor 
488  (1906) 

ZOTT,  63 

Zsigmondy,  R.,  5*,  6,  10,  41,  48-''",  49, 
49^",  73*2,  75^  75*2^  77^  83,  85,  92*3, 
93"'-,  98,  99,  122,  125,  143,  342 

1)  Liebig's  Ann.  301,  39  (1898) 

2)  Z.  Erkenntris  d.  KoU.  (Jena,  1905) 

3)  KoU.  Zeitschr.  8,  123  (1911) 
and  Schulz,  Fr.  N.,  143 

and  Siedentopf,  6,  75 
Wilke-Dörfürt  and  Galecki,  98 
Alexander's  translation,  247 

ZuNTZ,  N.,  216 

and  Schumberg,  216 

ZuNZ,  E.  and  Jacque,  L.,  199* 
Edgar,  86,  89^",  167*',  205,  363 
Bull,  de  la  Soc.  de  Sc.  med.  et  nat. 
de  Bruxelles  67,  178-179  (1909) 


SUBJECT   INDEX 


Abrin,  204 
Absorption,  316 

alimentary,  317 

of  exudates,  323 

of  fats,  247 

influence  of  protective  colloids  on, 
247 

mechanism  of,  320,  322 

parenteral,  323 

percutaneous,  324 

of  water  and  crystalloids,  318 
Acetic  acid,  422 
Acids,  as  fixatives,  421 
Acid,  poisoning  with,  301-310 
Adjective  dyeing,  430 
Adsorption,  19,  21,  22,  84,  109 

abnormal,  27 

affinitive  cm'ves,  25 

apparatus,  112 

arsenious  acid  in  iron  hydroxid  gel,  27 

change  induced  by,  109 

detergents,  28 

determination  of  electric  charge  by, 
112 

and  disinfectant  action,  398,  399,  400 

of  dyes,  25 

dyeing,  27 

effect   of   on  membranes    (see  Pro- 
teins), 58 

of  enzymes,  185 

equilibrium,  27-111 

gas  exchange,  309 

influence  of,  in  hemoglobin,  305,  306 

grapliic  of,  24 

in  microscopic  staining,  425 

and  immunity  reactions,  198 

influence   of    chemical    composition, 
27  et  seq. 

mechanical,  26 

of  narcotics,  389 

negative,  19-24 

negative,  simulation  of,  27 

pharmacological,  363 

reversible,  199 


Adsorption,  saturation,  26 

selective,  32 

specific,  198 

by  starches,  135 

therapy,  353,  et  seq. 

by  ultrafilters,  101 

and  urine  secretion,  337 
Age,  influence  of,  55 

influence  of,  on  meat,  169  et  seq. 
Agglutinin,  194,  195,  201,  202 
Albmnins,  146 

acid,  152 

alkali,  153 

amphoteric,  154 

coefficient  of  diffusion  of,  146 

electrolyte  free,  147,  156 

influence  of  inorganic  hydrosols  on, 
156 
Albumin,  in  milk,  349 

as  sols,  147 
Albuminoids,  161 
Albumoses,  166 
Alcohol,  as  fixative,  423 

effect  on  colloids  after  ingestion,  390 
Alcoholism,  324,  325 
Aluminimn,  382,  383 
Amboceptor,  195-200,  201,  206 
Ameba,  phagocytosis  by,  285 

migration  of,  284 
Anesthetics,  385 
Anaphylatoxin,  209 
Anaphylaxis,  209 

and  heavy  metals,  382 
Anion  and  cation  influence,  table,  297 

purgative  action  of,  412 
Antagonism  of  diuretics  and  narcotics, 

411 
Antagonism  of  ions,  82 

of  salts  physiological,  379 
Antibodies,  195-197 
Anti-enzymes,  191 

antigens  and  immune  substance,  205 
Antigens,  196 
Antitoxins,  191,  195 


457 


458 


SUBJECT  INDEX 


Anthrax,  disinfectant  action  on,  396, 
397 

inhibition  of,  growth  of,  407,  408 
Artefacts,  264 
Assimilation,  245 
Astringents,  414 

action  of,  383 
Avogadro's  law,  43 

Bacterial  staining,  435 
Balneology,  414 
Beer,  179 

cloudiness  of,  180 

fermentation  of,  181 

protective  colloids  of,  181 
Bile,  330 
Bio-colloids,  129 
Biological  determination  of  adsorption 

in  disinfective,  399 
Blood,  299 

reaction  of,  301 

corpuscles,  204,  244 

corpuscles,  303 

corpuscles,  composition  of,  304 

corpuscles,    influence    on   viscosity, 
314-315 

corpuscles,      hemolj^sis      (see      also 
Wasserman    reaction),    244 

corpuscles,  osmotic  pressure  of,  304 

corpuscles,  structure  of,  305 
Blotting    paper,    disinfectant     testing 

with,  408 
Boyle's  law,  51 
Bread,  177 

action  of  hemaglobin  in,  308  et  seq. 

gluten  restoration  of,  177 

staleness  of,  178 

war,  178 
Bromin,  therapeutic  action  of,  381 
Bronchial  glands,  328 
Brownian  Zsigmondy  movement,  49- 

53 
Bubble  method,  of  examining  milk,  173 
Buffer  substances  of  blood,  300 
Butter,  175-346 

Calcium,  action  of,  298A,  379-381 
compounds,  colloidal  preparations  of, 

381 
condition  of,  in  serum,  302 
ion,  influence  on  phagocytosis,  287 


Calcium,   phosphate,  condition  of,   in 
milk,  349 

utilization,  325,  361 
Calculi,  urinary,  343 
Carbohydrates,  133 
Carrel-Dakin  disinfection,  405 
Casein,  163,  348 
Cations  as  diuretics,  409 
Casts,  urinary,  344 

Carbon  dioxid,  influence   of,  on  m-ine 
excretion,  333 

solubility  in  blood,  309,  310 
Catalysers,  31,  183 
Catalysis,  81 
Cell,  276 

membrane,  279  et  seq. 

structure  of,  276 

cerebrospinal  fluid,  354 
Cheese,  176 
Chemical    attraction    of    precipitates, 

260 
Chemical  combination,  19,  22 

determination  of  adsorption  in  dis- 
infection, 398 

theory  of  dyeing,  425,  426 
Chemotaxis,  286 
Chloroform  poisoning,  delayed,  389 

distribution  of,  in  disinfection,  398 
Cholesterin,  87,  140,  141 
Chromic  acid,  421 
Circulation,  of  crystalloids,  235,  238 

coUoids,  239 

of  gases,  235 

of  material,  235 

of  water  (see  also  swelling),  236  et  seq. 
Clotting  of  blood,  300 
Cloudy  swelling,  228 
Coagulation,  82,  114,  142,  149 

of  blood,  effect  of  gelatin  on,  365 

chemical,  143,  149 

fractional,  82 

by  freezing,  144 

by  heat,  142,  151 

irreversible,  143 

by  light,  144 
Co-enzymes,  191 
Collagen,  161 
Colloidal  protection,  in  milk,  349 

in  urine,  343 
Colloids,  aging  of,  72,  74 

artificial,  5 


SUBJECT  INDEX 


459 


Colloids,  crystallization  of,  71 
color  of,  5,  7 
consistency  of,  64 
death  of,  73 
definition  of,  3 

dynamic  balance  in  organism,  308 
electrolytes,  46 
electrical  properties  of,  77 
electrical  production  of,  4 
hydrate,  36 
hydrophile,  9 
hydrophobe,  9 

importance  in  body,  126  et  seq. 
intravenous  action  of,  365 
life  curve  of,  72  et  seq. 
mechanical  production  of,  4 
migration  of,  84,  86 
optical  properties  of,  75 
particle  size,  7 

pharmaceutical  action  of,  362 
protective,  7,  11,  36,  77,  86,  181,  203, 

363 
protective,  in  urine,  343 
Colloid  swelling  and   urine  secretion, 
323,  336 
swelling,  and  equilibrium,  341 
■Colloidal  properties  of  dye  mixtures, 
430 
antimony,  377 
arsenic,  377 
mercury,  376 
phosphorus,  377 
metals,  therapeutic  use  of,  365 
silver,  366 

silver,  distribution  of,  373 
silver,  effect  on  blood,  371 
suver,  effect  on  temperature,  373 
silver,  therapeutics  of,  374 
silver,  in  infections,  376 
silver,  in  pneumonia,  375 
silver,  in  wounds,  366 
silver,  protective  colloids  for,  366 
sulphm-,  376 

swelling  state  of  tissues,  415 
Complement  deviation  or  fixation  (see 
also  Wassermann  reaction),    194, 
206,  207,  208 
Complement,  189,  198,  200,  206 
Concentration,  by  absorption,  27 

couples,  62 
■Conduction  by  nerves,  354 


Cooperation  of  drugs,  361 
Cream,  175,  346 

artificial,  175 

sophistication  of,  175 
Critical  narcotic  concentration,  385 
Cryoscopy,  341 
Crystals,  force  producing,  17 

Dehydrating  action  of  purgatives,  414 
Dehydration  of  food,  169 
Development,  252 
Diabetes  insipidus,  341 
Dialysis,  collodin  sac,  91 

methods,  89  et  seq. 
Diaphragms,  charge  of,  78 
Diarrhea,  321,  323 

treatment  of,  364 
Difl^usion,  103  et  seq. 

apparatus,  105,  106 

coefficient  of,  45,  52,  190 

influence  of  adsorption  upon,  55 

influence  of  substances  on,  55 

in  jellies,  54 

relation  to  dyeing,  428 

of  protein,  146 
DigestibUity  of  mük,  349,  350 
Digitalis,  411 

Disinfectants  and  dissociation,  402 
Disinfectant  action  of  cresol,  396 

action  of  H  and  OH  ions,  394 

action  of  phenyl  group  and  halogens, 
394 

action  of  sulpho  groups,  394 

and  adsorptive  capacity,  394,   395, 
396,  397 

action  of  chloroform,  398 

action  and  dilution,  396 

and  death,  396 

and  inhibition,  396 

of  specific  character,  394 

salts  of  heavy  metals,  394 
Disinfectants,  testing  oi,  405,  406 
Disinfection,  359 

definition  of,  391 

of  the  skin,  395 

mechanism  of,  391 

and  permeability,  402 

and  surface  tension,  391 
Dispersed  phase,  (see  Phase)  5,  11,  12 
Dispersion,  3 
Dissimilation,  245 


460 


SUBJECT  INDEX 


Dissimilation,  influence  of  enzymes,  250 
Dissociation  and  disinfection,  402 
Distribution,  20,  22,  31  et  seq. 

of  disinfectant,  398,  399 

of   disinfectant    on    microorganism, 
393 

Henry's  law,  20 

in  toxicology,  360 
Diuretics,  409 
Diuretic  action  of  chloral  caffeine,  362 

of  caffeine,  411 

of  salts  administered  intravenously, 
410 

of  theobromin,  411 

of  urea,  411 
Double  staining,  434 
Drugs,  influence  of,  on  kidneys,  338 
Dyeing  (see  Staining),  200-206,  407 

Eczema,  234 

Edema,  223  et  seq.,  377 

controversy  concerning,  229  et  seq. 

of  the  brain,  231,  352 

treatment  of,  339 
Elastic  fibers,  434 
Electric  charge,  enzymes,  187 

migration,  118 

migration,   apparatus  for   study  of, 
118 
Electrodes  —  nonpolarizable,  119 
Electro-endosmosis,  78 
Electrolyte  (see  also  Salt) 
Electrolytes,  149 

influence  of,  on  viscosity  of  gelatin, 
162 
Emulsion,  definition  of,  5 
Emulsions,  formation  of,  140 
Enzymes,  182 

adsorption  analysis  of,  185 

aging  of,  188 

colloidal  nature  of,  183 

diffusion  coefficient  of,  190 

electric  charge  of,  187 

inactivation  of,  189 

purification,  187 

specification  of,  188 

synthesis  by,  188 

ultrafiltration  of,  190 
Equilibrium,  341 

emulsion,  38 

irreversible,  28 


Equilibrium,  reversible,  28 

study  of  by  ultrafiltration,  102 
Erythrocytes     (see    Blood    corpuscles, 

volume  of),  307 
Excretion,  326 
Exudate,  223 

Fatty  degeneration,  nature  of,  377 
Fats,  deposition  of,  246 

resorption  of,  246 
Fibrin,  160 
Flocculation,  83,  86,  117 

vs.  salting  out,  83,  87 
Flour,  176 
Foods,  168 
Ferric  hydroxid,  negative  coUoid,  384 

positive  colloid,  384 
Ferric  oxid  as  arsenic  antidote,  385 

intravenous  injection  of,  384 

negative,  384 

positive,  384 
Fixatives,  420 
Fixing  and  hardening  of  tissues,  419^ 

420 
Formaldehyde,  distribution  of,  in  dis- 
infection, 398 

as  fixative,  423 
Freezing,  216 
Freezing-point  depression,  in  milk,  340 

in  urine,  351 
Friction  internal  (see  Viscosity),  64 

Gas  exchange  (see  Respiration) 
Gastric  juice,  329 
Gay-Lussac's  Law,  51 
Gel,  definition  of,  4,  8 

elastic,  66 

freezing  and  thawing  of,  66 
Gelatin,  161 
Gelatinization,  161  et  seq. 

time  of  for  agar,  138 
Gibbs'  Theorem,  25 
Glaciation,  216 
Gland,  326 
Glaucoma,  227 
Globulins,  158 

artificial,  159 
Gold  figure,  85 
Glycogen,  staining  of,  433 
Golgi's  stain,  mechanism  of,  431 
Gout,  148 


SUBJECT  INDEX 


461 


Gram's  stain,  30,  435 
Growth,  252 

biological,  with  shrinking,  265 
Growth  of  plants,  2()7 

influence  of  chemical  reaction,  267 

Hardening,  histological,  424 

Heavy  metals  and  salts  of,  157 

Heavy  metals,  as  £_>catives,  421 

Heavy  metal  specificity,  383 

Hematin,  165 

Hemoglobin,  164 

function    of,    309    (see    also    Blood 

corpuscles,  respiration) 
synthetic,  385 

Hemolysis,  305,  306 
induced  by  colloids,  371 

Hemolysins,  196,  200 

Honey,  176 

H  ion  concentration  in  blood,  302 
influence  upon  circulation,  314 
influence  upon  erythrocytes,  312 
influence  of  CO2  upon,  311 

Histone,  160 

Hydremia,  341 

Hydrosol,  11 

Immune  substances,  201 
Immunity  reactions  (see  also  Precipi- 
tin), 193  et  seq. 
Inactivation,  by  shaking,  34 
Inflammation,  232 
Inhibition,  354 

zones  or  irregular  series,  84,  118,  140 
Instant  values,  51 
Interferometer,  120 
Interface  (see  Surface) 
Interface,  14 
Internal  friction,  113 

of  albumin,  152 

of  globulin,  159 

of  gums,  137 
Intestinal  inflammation,  322. 
Intestinal  secretion,  330 
Inulin,  133,  210 

lodin,  therapeutic  action  of,  380 
Iron,  astringent  action,  383 

as  arsenic  antidote,  384 

action  ot  colloidal,  383 

colloidal  preparations,  value  of,  383, 
384 


Iron,  mechanism  in  hemostasis,  384 
intravenous  injection  of,  384 
oral  administration  of,  384 
pharmacological  action  of,  383 

Irregular  series,  157,  203 

Irritability  of  nerves,  353 

Isoelectric  zone,  77,  84,  161 
point  for  casein,  164 
hemoglobin,  165 

Jellies,  structure  of,  9 

Keratins,  163 
Kinetic  theory,  50 

Lanolin,  composition  of,  416 

properties  of,  416 
Layered  structm-es,  261  et  seq. 
Lecithin,  87,  139,  140 

precipitation  of  magnesium  salts,  388 
Liesegang' s  rings,  263 
Lipoids,  definition  of,  139 

reaction  with  narcotics,  389 

staining  of,  433 
Local  anesthetics,  mechanism  of,  389 
Lymph,  303 
Lyotropic  series,  140 
Lyotropism,  definition,  81 

Maceration  for  microscopic  study,  419 
Magnesium  sulphate,  pm-gative  action 

of,  413 
Margarine,  175 
Mass  staining,  431 
Meat,  169 

boiling,  172 

cold  storage,  169 

preserving,  172 
Meiostagmin  reaction,  211 
Melting  temperature,  114,  138,  161 
Membrane,  definition  of,  56 

formation  of,  56 

growth,  57 

self  regulation,  58 

semipermeable,  57,  236 

and  crystalloids,  58 

equilibria,  59 

hydrolysis,  59 

and  substances  interchange,  239 
Membranes,  mfluence  on  substance  in- 
terchange, 239 


462 


SUBJECT  INDEX 


Membranes  and  electrolytes,  59 
Mercuric  chlorid  poisoning,  143 
Mercury  poisoning,  383 

salts,  as  disinfectant,  400 
Metamorphosis,  252 
Microorganisms,   influence  of   suspen- 
sions on  growth,  393 
Microscopic  technic,  417 
Milk,  adulteration  of,  174 

colloids  of,  174,  349 

condensed,  174 

cows  and  woman's,  349,  351 

dried,  351 

food  accessories  of  McCollum,  351 

freezing  point  depression  of,  351 

homogenized,  346,  348 

raw  and  boiled,  350 

surface  pellicles  in,  346 

skin  formation  in,  350 

ultrafiltration  of,  350 

ultramicroscopy  of,  349 

viscosity  of,  173,  348 

drying  of,  174 

examination  of,  173 

ultrafiltration  of,  174 
Molecule,  42 
Molecular,  movement,  50 

weight,  42  et  seq. 
Mordants,  431,  434 
Movement  of  organisms,  282 
Myosin,  160 

Mucins  and  mucoids,  165 
Muscle,  289  et  seq. 

influence  of  Ca  on,  298a 
Muscles,  influence  of  drugs  on,  298a 

influence  of  lactic  acid  on,  298 

electric  phenomena  of,  294 

fatigue,  291 

function,  292,  296,  298 

influence  of  electrolytes  on,  294 

influence  of  phosphates  on,  298a 

working  model  of,  296 

Narcosis,  distribution  of  narcotic,  388 

and  oxygen  absorption,  388 

and  respiration,  388 

and  change  in  turgor,  387 
Narcotics,  385 

action  and  electric  conductivity,  386 
-  action  and  inhibition  of  hemolysis, 
386 


Narcotics,  action  of,  on  permeation  of 
electrolytes,  389 
effect  of,  on  plasma  pellicle,  389 
lipoid  solution  theory,  385 
Meyer-Overton  theory,  385 

Narcotics,  toxic  action  of,  390 

and  disturbance  of  oxidation,  390 
action  on  catalase,  390 

Narcotic  action  and  inhibition  of  fer- 
ment action,  386 
and  inhibition  of  oxidation,  386 
action  and  lipoid  solubility,  386 
action  of  magnesium  salts,  386,  388 
action  and  protein  precipitation,  386 
action  and  plasma  pellicle,  386 
action  and  surface  tension,  386 

Nephelometer,  120 

Nephritis,  338 

Nerves,  352 

Neurobiotaxis,  261 

Neutral  salts  toxic  action,  379 

Nucleins,  160 

Nucleoalbumins,  163 

Nucleus,  267,  279 
staining  of,  433 

Organism,  circulation  of,  23 

as  colloid,  213 
Optical  methods,  119 

rotation,  156 
Osmic  acid,  421 

Osmotic  compensation  method,  appifeCi 
to  milk,  348 

growths,  250 

pressure,  measurement  of,  43,  51,  10 1 

instruments,  106 

compensation  method,  107 

measurements  of  starches,  135 
Ossification,  268 

theories  of,  269 
Oxygen  and  air  as  disinfectants,  399 

Pancreatic  juice,  330 
Parthenogenesis,  245 
Particle  size  (see  also  Proteins),  41 
Pellicle,  plasma,  nature  of,  240  et  seq. 

of  blood  corpuscles,  301 
Peptisation,  84 

Permeability,  of  cell  membrane,  242, 
243 

chemical  regulation  of,  244 


SUBJECT  INDEX 


463 


Permeability,  lethal  change  of,  245 

photo  regulation  of,  245 

selective,  63 
Peptones,  166 
Phagocytosis,  285,  287  et  seq. 

induced  by  colloidal  silver,  372 

influence  of  cation,  287 
Phase,  definition  of,  5 

influence  of  serum,  288 
Phenol,  distribution  of,  in  disinfection, 

398 
Photodromy,  76 
Picric  acid,  422 

after  vital  staining,  422 
Plasma,  299 
Plasma  pellicle,  effect  of  narcotics  on, 

389 
Plasmolysis,   of    aspergillus    cells    (see 

also  Hemolysis),  241,  243 
Poisons,  360 

Powders,  therapeutic  effect  of,  364 
Precipitation,  80  et  seq.,  157  (see  also 
Coagulation) 

by  alcohol,  152 

influence  of  electrolytes,  163 

irreversible,  87 

of  gelatin,  162 

zones,  157 
Precipitin  reaction,  56 

(see  also  Immunity  reaction,  201, 202) 
Pro-enzymes,  191 
Protamine,  160 
Protection,  preferential,  371 
Protective  ferment,  210 
Proteins,  142 

adsorption  phenomena,  145  et  seq. 

crystallization  of,  144 

ultrafiltration  of,  146 

ultramicroscopic  studies  of,  144 
Proteoses,  167 
Protoplasm,  277  et  seq. 

death  of,  279 

staining  of,  433 
Pulsation,   influence  of,   on  secretion, 

327,  332 
Purgatives,  409 

mechanism  of,  412 
Pyrosol,  11 

Radioactivity,  therapeutic  action,  415 
Radioactive  substances,  87 


Regulation,   automatic  of   cell   metab- 
olism, 244 
Respiration  (see  Gas  exchange) 
Rhythmic  phenomena,  263 
Ricin,  204 
Rigor  Mortis,  291 

Saccharo-coUoids,  133 

Saliva,  328 

Salts,  concentration  of  in  intestines,  412 

Salt,  distribution  of,  233 

Salting  out,  80 

Saponin,  34 

Salts  as  fixative,  422 

therapeutic  and  toxic  action  of,  378 
Salves,  mechanism  of,  416 
Secretion,  315  ei  seq. 
Section  staining,  431 
Separation,  by  shaking  out  the  foam,  35 
Serum,  content  of  globulin,  159 

influence  of  on  solubility  of  salts,  302 

surface  tension  of,  303 
Shrinking,  65 

Silicic  acid  jelly,  size  of  pores,  10 
Silk  threads,  disinfectant  testing  with, 

406 
Silver  nitrate,   disinfectant   action  of, 

39S 
Size,  therapeutic,  use  of,  365 
Skin  disinfection,  404 
Soap  as  a  disinfectant,  404 
Sol,  definition  of,  3 
Solidification  temperature,  114,  161 
Solubuity,  influence  of  gelatin  on,  161 

selective,  63 
Solutions,  homogeneous,  6,  19 
Specific  action  of  disinfectants,  401,  402 
Specific  chemical  action  of  disinfectant, 

401 
Spongin,  163 
Staining,  424 

bacterial,  435 

double,  434 

method  of,  431  et  seq. 

technic  of,  431 

theory  of,  424  et  seq. 
Starch,  133 

agar,  136 

cellulose,  138 

crystaUizable,  135 

glucosides,  136 


464 


SUBJECT  INDEX 


Starch,    granules,   relation  to  mineral 
content,  134 

gums,  136 

soluble,  134 

paste,  134,  135 

molecule,  size  of,  135 
Structures,  genesis  of,  259 

osmotic,  253  et  seq. 
Surface,  development  of,  13 

phenomena  of,  11 

pellicles,  in  milk,  346 

tension,  287 

tension,  and  disinfection,  391 

tension,  measiu-ement  of,  108 

tension,  and  muscular  action,  296 

tension,  by  ultrafiltration,  16 
Surface  tension,  14,  81 

in  common  things,  34 

skins,  33 

of  solids,  18 

in  stains,  34 

in  milk,  173 
Suspension,  definition  of,  5 
Sweat  glands,  345 
Swelling,  65,  169,  173 

influence  ci  electrolytes  on,  67  et  seq., 
222 

and  intestinal  absorption,  320 

of  gelatin,  161 

of  organs,  217 

measurements  of,  114 
•  pressure,  66,  115 

range,  217,  219 

ratio,  217 

Threshold  "electrolyte,"  83,  118 
Tissue  behavior  with  dyes  and  fixa- 
tives, 433 
growth  in  vitro,  246 
Tropisms,  283 
Toxin-antitoxin,  31 
Tropisms,  explanation  of,  283 
Turgor  and  irritability,  387 
Tyndall  phenomenon,  75 

Ultracentrifugation,  6,  42 
Ultrafilter,  theory  of  dyeing,  427 

theory  of  plasma  pellicle,  241 
Ultrafiltration,  6,  10,  42,  58 

absorption  in,  101 

applications  of,  102 


Ultrafiltration,  gauging  of,  99 

methods  of,  and  apparatus,  95  et  seq. 

of  albumoses,  166 

of  cerebrospinal  fluid,  354 

of  enzymes,  190 

of  milk,  174 

of  proteins,  144 

of  viruses,  393 
Ultramicroscope,  6,  75,  122  et  seq. 

cardioid  condenser,  126 
Ultramicroscopy  of  milk,  349 
Urea,  diuretic  action  of,  334  • 

influence  on  diffusion,  55 
Ultrafiltration,  327,  332 

of  urine,  332 
Urine,  colloids  of,  342 

concentration  of,  335,  336 

effect  of  drugs  on,  338 

freezing  point  depression,  340 

glomerular  secretion  of,  331  et  seq. 

normal,  342 

pathological  excretion  of,  338,  343 

secretion  of,  323,  330,  332,  336 

surface  tension  of,  343,  344 

threshold,  substances  in,  336 

Viscosity,  113 

of  albumins,  151,  154 

of  blood,  310,  311,  312 

of  blood  and  diuresis,  334 
Viscosity  of  plasma,  311 
Viscosimeters,  113 
Viscosity,  influence  of  caffeine  on,  152 

influence  of  neutral  salts  on,  152 
Viscosity  of  serum,  302 
Vital  staining,  431 

Wassermann    reaction     (see     Comple- 
ment deviation) 
Water,  in  blood,  220 

content  influence  on  muscle  function, 
298 

distribution  in  body,  415 

distribution  of,  in  organism,  215 

in  muscles,  220 

in  organs  (see  Swelling),  217,  218, 
219,  221 

pathology  of,  223 

of  solution,  66 

of  swelling,  66 
Wood  formation,  248  et  seq. 
Wound  disinfection,  404,  405 


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Casler,  M.  D.     Simplified  Reinforced  Concrete  Mathematics i2mo,  ^^i  00 

Cathcart,  W.  L.     Machine  Design.     Part  I.  Fastenings 8vo,  *3  00 

Cathcart,  W.  L.,  and  Chaffee,  J.  I.     Elements  of  Graphic  Statics .  .  .8vo,  *3  00 

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Caven,  R.  M.,  and  Lander,  G.  D.    Systematic  Inorganic  Chemistry.  i2mo,  2  25 

Chalkley,  A.  P.     Diesel  Engines 8vo,  *4  00 

Chalmers.  T.  W.     The  Production  and  Treatment  of  Vegetable  Oils, 

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Chambers'    Mathematical    Tables 8vo,  2  50 

Chambers,  G.  F.     Astronomy i6mo,  *i  50 

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Chatley,  H.     Principles  and  Designs  of  Aeroplanes i6mo,  o  75 

How  to  Use  Water  Power i2mo,  *i  50 

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Child,  C.  D.     Electric  Arc 8vo,  *2  00 

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Clapperton,   G.     Practical    Papermaking 8vo    (Reprinting.) 

Clark,  A.  G.     Motor  Car  Engineering. 

Vol.    I.     Construction *4  00 

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Clark,  C.  H.     Marine  Gas  Engines.     New  Edition 2  00 

Clarke,  J.  W.,  and  Scott,  W.    Plumbing  Practice. 

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Clarkson,  R.  P.     Elementary  Electrical  Engineering  (In  Press.) 

Clerk,  D.,  and  Idell,  F.  E.    Theory  of  the  Gas  Engine i6mo,      o  75 

Clevenger,  S.  R.     Treatise  on  the  Method  of  Government  Surveying. 

i6mo,    morocco, 

Clouth,  F.     Rubber,  Gutta-Percha,  and  Balata 8vo, 

Cochran,  J.     Concrete  and  Reinforced  Concrete  Specifications 8vo, 

■ Treatise  on  Cement  Specifications 8vo, 

Cocking,  W.  C.     Calculations  for  Steel-Frame  Structures i2mo. 

Coffin,  J.  H.  C.     Navigation  and  Nautical  Astronomy lamo, 

Colbum,  Z.,  and  Thurston,  R.  H.     Steam  Boiler  Explosions.  ..  .i6mo, 
Cole,   R.   S.     Treatise   on   Photographic   Optics lamo, 

Coles-Finch,  W.     "Water,  Its  Origin  and  Use 8vo, 

Collins,  C.  D.    Drafting  Room  Methods,  Standards  and  Forms 8vo, 

Collins,  S.  Hoare.     Plant  Products  and  Chemical  Fertilizers 8vo, 

Collis,  A.  G.     High  and  Low  Tension  Switch-Gear  Design 8vo, 

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Comstock,  D.  F.,  and  Troland,  L.  T.     The  Nature  of  Electricity  and 

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Cowell,  W.  B.     Pure  Air,  Ozone,  and  Water r2mo, 

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Crocker,  F.  B.,  and  Wheeler,  S.  S.  The  Management  of  Electrical  Ma- 
chinery   i2mo,     *i  00 

Crosby,  E.  U.,  Fiske,  H.  A.,  and  Forster,  H.  W.     Handbook  of  Fire 

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2 

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Crosskey,  L.  R.,  and  Thaw,  J.     Advanced  Perspective.... 8vo,  2  00 

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Darling,  E.  R.     Inorganic   Chemical  Synonyms iimo,  i  00 

Davenport,  C.     The   Book 8vo,  2  00 

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Davies,  F.  H.    Electric  Power  and  Traction 8vo,  *2  00 

— — Foundations   and   Machinery   Fixing i6mo,  i  00 

Deerr,   N.     Sugar   Cane 8vo,  10  00 

De  la  Coux,  H.    The  Industrial  Uses  of  Water 8vo,  5  00 

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Denny,  G.  A.     Deep-level  Mines  of  the  Rand 4to,  *io  00 

De   Roos,  J.  D.  C.     Linkages lömo,  o  75 

Derr,  W.  L.     Block  Signal  Operation Oblong  i2mo,  *i  50 

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De  Varona,  A.     Sewer  Gases .  .    i6mo,  075 

Devey,  R.  G.     Mill  and  Factory  Wiring i2mo,  i  00 

Dichmann,   Carl.     Basic   Open- Hearth   Steel    Process i2mo,  4  00 

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Dilworth,  E.  C.     Steel  Railway  Bridges 4to.  '''4  00 

Dinger,  Lieut.  H.  C.     Care  and  Operation  of  Naval  Machinery.  ..  i2mo,  *3  00 
Dixon,  D.  B.     Machinist's  and  Steam  Engineer's  Practical  Calculator. 

lömo,  morocco,  i  25 

Dommett,   W.   E.     Motor  Car  Mechanism i2mo,  '""'a  00 

Dorr,  B.  F.     The  Surveyor's  Guide  and  Pocket  Table-book. 

löino,  morocco,  2  00 

Draper,  C.  H.     Heat  and  the   Principles  of  Thermo-Dynamics.  .i2mo,  2  25 

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Dubbel,  H.     High  Power  Gas  Engines 8vo,  *5  00 

Dumesny,  P.,  and  Noyer,  J.     Wood  Products,  Distillates,  and  Extracts. 

8vo,  *5  00 
Duncan,  W.  G.,  and  Penman,  D.     The  Electrical  Equipment  of  Collieries. 

8vo,  *5  00 

Dunkley,  W.  G.   Design  of  Machine  Elements.   Two  volumes.  .8vo,each,  2  00 

Dunstan,  A.  E.,  and  Thole,  F.  B.  T.     Textbook  of  Practical  Chemistry. 

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Durham,  H.  W.     Saws .  Svo,  2  50 

Duthie,  A.  L.     Decorative  Glass  Processes Svo,  2  50 

Dwight,  H.  B.     Transmission  Line  Formulas Svo,  *2  00 

Dyke,  A.  L.     Dyke's  Automobile  and  Gasoline  Engine   Encyclopedia, 

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Dyson,  S.  S.     A  Manual  of  Chemical  Plant.     12  parts.  ..  .410,  paper,  7  50 

Dyson,  S.  S.,   and  Clarkson,  S.  S.     Chemical  Woiks 8vo,  ^9  00 

Eccles,  W.  H.     Wireless  Telegraphy  and   Telephony i2mo,  *8  80 


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Eck,  J.     Light,   Radiation   and  Illumination 8vo, 

Eddy,  L.  C.     Laboratory  Manual  of  Alternating  Currents lamo, 

Edelman,  P.  Inventions  and  Patents i2mo, 

Edgcumbe,  K.     Industrial  Electrical  Measuring  Instruments 8vo, 

Edler,    R.      Switches    and    Switchgear Svo, 

Eissler,  M.     The  Metallurgy  of  Gold 8vo, 

The  Metallurgy  of  Silver 8vo, 

The   Metallurgy   of   Argentiferous    Lead Svo, 

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Ekin,  T.  C.     Water  Pipe  and  Sewage  Discharge  Diagrams folio, 

Electric  Light  Carbons,  Manufacture  of Svo, 

Eliot,  C.  W.,  and  Storer,  F.  H.     Compendious  Manual  of  Qualitative 

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Ennis,  Wm.  D.     Linseed  Oil  and  Other  Seed  Oils 8vo, 

■ Applied    Thermodynamics 8vo, 

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Vapors  for  Heat  Engines i2mo, 

Ermen,  W.  F.  A.     Materials  Used  in  Sizing 8vo, 

Erwin,  M.     The  universe  and  the  Atom i2mo   (Reprinting.) 

Ewing,  A.  J.     Magnetic  Induction   in  Iron 8vo,      5  00 

Fairchild,  J.  F.     Graphical  Compass  Conversion  Chart  and  Tables...  o  50 

Fairie,  J.     Notes  on  Lead  Ores izmo,  *o  50 

Notes  on  Pottery  Clays i2mo,  *2  00 

Fairley,  W.,  and  Andre,  Geo.  J.     Ventilation  of  Coal  Mines.  ..  .i6mo,  o  75 

Fairweather,  W.  C.     Foreign  and  Colonial  Patent  Laws 8vo,  *3  00 

Falk,   K.    G.     Chemical    Reactions:    Their    Theory   and   Mechanism. 

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Fanning,  J.  T.     Hydraulic  and  Water-supply  Engineering 8vo,  *S  00 

Farnsworth,  P.  V.     Shop  Mathematics.  . .         i2mo  (In  Press.) 

Fay,  I.  W.     The  Coal-tar  Dyes 8vo,  5  oa 

Fernbach,  R.  L.     Glue  and  Gelatine , 8vo,  *3  00 

Findlaj^  A.    The  Treasures  of  Coal  Tar i2mo,  2  00 

Firth,  J.  B.    Practical  Physical  Chemistry i2mo,  i  25 

Fischer,  E.     The  Preparation  of  Organic  Compounds T2mo,  i  50 

Fisher,  H.  K.  C,  and  Darby,  W.  C.     Submarine  Cable  Testing.  .  ,8vo,  4  00 

Fleischmann,  W.     The  Book  of  the  Dairy Svo,  4  50 

Fleming,  J.  A.     The  Alternate-current  Transformer.     Two  Volumes.  8vo. 

Vol.     I.     The  Induction  of  Electric  Currents *6  50 

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Propagation    of    Electric    Currents 8vo,  3  50 

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Fleur}',  P.     Preparation  and  Uses  of  White  Zinc  Paints 8vo,  3  00 

Flynn,  P.  J.     Flow  of  Water i2mo,  o  75 

Hydraulic    Tables r6mo,  o  75 


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Foye,  J.  C.     Chemical  Problems lömo, 

Handbook    of    Mineralogy i6mo, 

Francis,  J.  B.     Lowell  Hydraulic  Experiments 4to, 

Franzen,  H.     Exercises  in  Gas  Analysis i2mo, 

Fräser,   E.   S.,   and  Jones,   R.   B.     Motor   Vehicles   and  Their  Motors, 

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Freudemacher,  P.  W.     Electric  Mining  Installations .lamo. 

Friend,  J.  N.     The  Chemistry  of  Linseed  Oil i2mo, 

Fritsch,  J.     Manufacture  of  Chemical  Manures 8vo, 

Frye,  A.  I.     Civil  Engineers'  Pocket-book i2mo,  leather, 

Fuller,  G.  W.     Investigations  into  the  Purification  of  the  Ohio  River. 

4to, 
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Gant,  L.  W.     Elements  of  Electric  Traction .8vo,     *2  50 

Garcia,  A.  J.  R.  V.     Spanish-English  Railway  Terms 8vo,       3  00 

Gardner,  H.   A.     Paint  Researches,  and  Their  Practical  Applications, 

8vo,    *5  00 
Garforth,  W.  E.     Rules  for  Recovering  Coal  Mines  after  Explosions  and 

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Garrard,  C.  C.     Electric  Switch  and  Controlling  Gear 8vo, 

Gaudard,    J.      Foundations i6mo, 

Gear,  H.  B.,  and  Williams,  P.  F.     Electric  Central  Station  Distribution 

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Geerligs,  H.  Ci  P.     Cane  Sugar  and  Its  Manufacture 8vo, 

Chemical  Control   m  Cane   Sugar  Factories 4to, 

Geikie,  J.     Structural  and  Field  Geology 8vo, 

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The  Antiquity  of  Man  in  Europe 8vo, 

Georgi,  F.,  and  Schubert,  A.     Sheet  Metal  Working 8vo, 

Gerhard,  W.  P.     Sanitation,  Watersupply  and  Sewage  Disposal  of  Country 

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Gas  Lighting    i6mo, 

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House    Drainage    i6mo, 

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Gibbings,  A.   H.     Oil   Fuel  Equipment   for  Locomotives.     8vo. 

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Gibson,  A.  H.     Hydraulics  and  Its  Application 8vo, 

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Gibson,  A.  H.,  and  Ritchie,  E.  G.    Circular  Arc  Bow  Girder ^to, 

Gilbreth,  F.  B.     Motion  Study i2mo, 

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Gillmore,  Gen.  Q.  A.    Roads,  Streets,  and  Pavements i2mo, 

Godfrey,  E.     Tables  for  Structural  Engineers i6mo,  leather, 

Golding,  H.  A.     The  Theta-Phi  Diagram i2mo, 

Goldschmidt,  R.     Alternating  Current  Commutator  Motor 8vo, 

Goodchild,  W.     Precious    Stones 8vo, 

Goodell,    J.    M.      The    Location,    Construction    and    Maintenance    of 

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Gore,  G.     Electrolytic  Separation  of  Metals 8vo, 

Gould,  E.  S.     Arithmetic  of  the  Steam-engine i2mo, 

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Gould,  E.  S.    Practical  Hydrostatics  and  Hydrostatic  Formulas.  .i6mo, 

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Greenwood,  H.    C.     The   Industrial   Gases 8vo    (In    Press.) 

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Grierson,  R.     Some  Modern  Methods  of  Ventilation 8vo, 

Griffiths,  A.  B.     A  Treatise  on  Manures lamo    (Rcprinliiu/.) 

Gross,  E.     Hops Svo, 

Grossman,  J.     Ammonia  and  Its  Compounds i2nio, 

Groth,   L.   A.     Welding  and  Cutting  Metals  by   Gases   or  Electricity. 

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Guldner,  H.     Internal  Combustion  Engines (/;;    Press.) 

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Guy,  A.  E.     Experiments  on  the  Flexure  of  Beams Svo, 

Haenig,   A.     Emery    and   Emery   Industry Svo, 

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Hale,    A.   J.      The   Manufacture    of    Chemicals    by    Electrolysis.     8vo, 

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Hall,  C.  H.     Chemistry  of  Paints  and  Paint  Vehicles i2mo, 

Hall,   R.   H.     Governors   and   Governing   Mechanism i2mo. 

Hall,  W.  S.    Elements  of  the  Differential  and  Integral  Calculus.  . .  .8vo, 
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Haller,  G.  F.,  and  Cunningham,  E.  T.     The  Tesla  Coil i2mo, 

Halsey,  F.  A.     Slide  Valve  Gears i2rao, 

The    Use    of    the    Slide    Rules i6mo, 

Worm   and    Spiral   Gearing T5mo, 


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12        D.  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG 

Hancock,  H.     Textbook  of  Mechanics  and  Hydrostatics 8vo, 

Hardy,  E.     Elementary  Principles  of  Graphic  Statics i2mo, 

Haring,  H.     Engineering  Law. 

Vol.  I.     Law  of  Contract 8vo, 

Harper,  J.  H.     Hydraulic  Tables  on  the  Flow  of  Water i6nio, 

Harris,  S.  M.     Practical  Topographical  Surveying {In  Press.) 

Harrow,  B.     Eminent  Chemists  of  Our  Times:  Their  Lives  and  Work. 

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Haskins,  C.  H.     The  Galvanometer  and  Its  Uses i6mo, 

Hatt,  J.  A.  H.     The  Colorist square  i2mo, 

Hausbrand,  E.    Drying  by  Means  of  Air  and  Steam lamo, 

Evaporating,  Condensing  and  Cooling  Apparatus , .  .8vo, 

Hausmann,   E.     Telegraph  Engineering .8vo, 

Hausner,  A.     Manufacture  of  Preserved  Foods  and  Sweetmeats.  ..  .Svo, 
Hawkesworth,  J.     Graphical  Handbook  for  Reinforced  Concrete  Design. 

4to, 

Hay,  A.     Continuous  Current  Engineering Svo, 

Hayes,  H.  V.    Public  Utilities,  Their  Cost  New  and  Depreciation. .  .8vo, 

Public  utilities,  Their  Fair  Present  Value  and  Return 8vo, 

Heath,  F.  H.    Chemistry  of  Photography 8vo.  {In  Press.) 

Heather,  H.  J.   S.     Electrical   Engineering 8vo,      450 

Heaviside,  0.     Electromagnetic  Theory.     Vols.  I  and  II....8vo,  each, 

{Reprinting.) 

Vol.    Ill 8vo     {Reprinting.)- 

Heck,  R.  C.  H.     The  Steam  Engine  and  the  Turbine 8vo,      4  50 

Steam-Engine  and  Other  Steam  Motors.     Two  Volumes. 

Vol.  I.     Thermodynamics  and  the   Mechanics 8vo,      4  50 

Vol.   II.     Form,   Construction,   and   Working 8vo,      5  50 

Notes  on  Elementary  Kinematics 8vo,  boards,     *i  00 

Graphics  of  Machine  Forces 8vo,  boards,     *i  00 

Heermann,   P.     Dyers'   Materials i2mo,      3  00 

Hellot,  Macquer  and  D'Apligny.   Art  of  Dyeing  Wool,  Silk  and  Cotton.  Svo,     *2  00 
Hering,  C,  and  Getman,  F.  H.     Standard  Tables  of  Electro-Chemical 

Equivalents    i2mo,    *2  00 

Hering,  D.  W.     Essentials  of  Physics  for  College  Students 8vo,      2  25 

Herington,  C.  F.     Powdered  Coal  as  Fuel Svo,      3  00 

Herrmann,  G.     The  Graphical  Statics  of  Mechanism i2mo,      2  00 

Herzfeld,   J.     Testing   of   Yarns   and    Textile    Fabrics Svo. 

{New  Edition  in  Preparation.) 

Hildenbrand,    B.    W.      Cable-Making i6mo,      o  75 

Hilditch,  T.  P.     A  Concise  History  of  Chemistry i2mo,    *i  50 

Hill,  M.  J.  M.     The  Theory  of  Proportion 8vo, 

Hillhouse,  P.  A.     Ship  Stability  and  Trim 8vo, 

Hiroi,  I.     Plate  Girder  Construction i6mo, 

>  Statically-Indeterminate  Stresses i2mo, 

Hirshfeld,  C.  F.    Engineering  Thermodynamics i6mo. 

Hoar,  A.     The  Submarine  Torpedo  Boat i2mo, 

Hobart,  H.  M.    Heavy  Electrical  Engineering 8vo, 

Design   of   Static   Transformers i2mo, 

Electricity 8vo, 

Electric  Trains 8vo, 

Electric  Propulsion  of  Ships 8vo, 


*2 

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50 

D.  VAN  xNOSTRAND  CO.'S  SHORT  TITLE  CATALOG        13 

Hobart,  J.  F.     Hard  Soldering,  Soft  Soldering  and  Brazing lamo,  1  25 

Hobbs,  W.  R.  P.    The  Arithmetic  of  Electrical  Measurements.  ..  .i2mo,  o  75 

Hoff,  J.  N.     Paint  and  Varnish  Facts  and  Formulas i2mo,  i  50 

Hole,  W.     The  Distribution  of  Gas 8vo,  *8  50 

Hopkins,  N.  M.     Model  Engines  and  Small  Boats i2mo,  i  25 

The    Outlook    for    Research    and    Invention i2mo.  2  00 

Hopkinson,  J.,  Shoolbred,  J.  N.,  and  Day,  R.  E.     Dynamic  Electricity. 

i6mo.  o  7"^ 

Horner,  J.     Practical  Ironfounding 8vo,  *2  00 

G«ar  Cutting,   in   Theory   and   Practice 8vo    {Reprinting.) 

Houghton,  C.  E.     The  Elements  of  Mechanics  of  Materials lamo,  2  50 

Houstoun,  R.  A.    Studies  in  Light  Production i2mo,  2  00 

Hovenden,  F.     Practical  Mathematics  for  Young  Engineers i2mo,  *i  50 

Howe,  G.     Mathematics  for  the  Practical  Man i2mo,  i  50 

Howorth,  J.     Repairing  and  Riveting  Glass,  China  and  Earthenware. 

8vo,  paper,  *o  50 

Hoyt,  W.  E.     Chemistry  by  Experimentation 8vo,  *o  70 

Hubbard,    E.      The    Utilization    of    Wood- waste Bvo,  *2  50 

Hübner,  J.    Bleaching  and  Dyeing  of  Vegetable  and  Fibrous  Materials. 

8vo   {Reprinting.) 

Hudson,  O.  F.     Iron  and  Steel 8vo,  ^  ^o 

Humphreys,  A.  C.  The  Business  Features  of  Engineering  Practice . 8vo,  2  50 

Hunter,  A.    Bridge  Work 8vo.  {In  Press.) 

Hurst,  G.  H.     Handbook  of  the  Theory  of  Color 8vo,  *3  5° 

Dictionary  of  Chemicals  and  Raw  Products 8vo,  *5  00 

Lubricating   Oils,  Fats   and  Greases 8vo,  *5  00 

Soapa    8vD,  *6  00 

Hurst,  G.  H.,  and  Simmons,  W.  H.     Textile  Soaps  and  Oils 8vo,  3  50 

Hurst,  H.  E.,  and  Lattey,  R.  T.    Text-book  of  Physics Svo,  *3  00 

Also  published  in  three  parts. 

Part      I.     Dynamics  and  Heat i  So 

Part    IL     Sound   and    Light i  5» 

Part  III.    Magnetism  and  Electricity *i  5» 

Hutchinson,  R.  W.,  Jr.     Long  Distance  Electric  Power  Transmission. 

i2mo,  *3  00 
Hutchinson,  R.  W.,  Jr.,  and  Thomas,  W.  A.    Electricity  in  Mining.  i2mo, 

{In  Press.) 

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Eyde,  F.  S.    Solvents,  Oils,  Gums,  Waxes 8vo, 

Induction  Coils    i6mo. 

Ingham,  A.  E.    Gearing.    A  practical  treatise 8vo, 

Ingle,  H.     Manual   of  Agricultural   Chemistry Bvo    {In   Press.' 

Inness,  C.  H.    Problems  in  Machine  Design i2mo, 

CeQtrifogal  Pumps   i2mo, 

The  Fan  i2mo, 

Jacob,  A.,  and  Gould,  E.  S.    On  the  Designing  and  Construction  of 

Storage   Reservoirs    i6mo.      o  75 

Jacobs,  F.  B.     Cam  Design  and  Manufacture (  In  Press.) 


0 

75 

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D.  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG 


James,  F.  D.     Controllers  for  Electric  Motors 8vo.      ^  oo 

Jehl,  F.     Manufacture  of  Carbons 8vo.      .■;  oo 

Jennings,  A.  S.     Commercial  Paints  and  Painting 8vo.      2  so 

Jennison,  F.  H.    The  Manufacture  of  Lake  Pigments.  .Bvo  {In  Press.) 

Jepson,  G.     Cams  and  the  Principles  of  their  Construction 8vo,     *i  50 

Mechanical  Drawing 8vo  {In  Preparation.) 

Jervis-Smith,   F.   J.     Dynamometers Bvo.      4.  00 

Jockin,  W.     Arithmetic  of  the  Gold  and  Silversmith i2mo,     *i  00 

Johnson,  C.  H.,  and  Earle,  R.   P.     Practical  Tests  for  the   Electrical 

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Johnson,  J.  H.     Arc  Lamps  and  Accessory  Apparatus i2mo,      o  75 

Johnson,  T.  M.     Ship  Wiring  and  Fitting i2mo    (Reprinting.)', 

Johnston,  J.  F.  W.,  and  Cameron,  C.     Elements  of  Agricultural  Chemistry 

and  Geology - izmo,      2  60 

Joly,  J.     Radioactivity  and  Geology i2mo    {Reprinting.) 

Jones,  H.  C.    Electrical  Nature  of  Matter  and  Radioactivity i2mo, 

Nature  of  Solution Bvo, 

. New  Era  in  Chemistry lamo, 

Jones,  J.  H.     Tinplate  Industry Bvo, 

Jones,  M.  W.    Testing  Raw  Materials  Used  in  Paint lamo, 

Jordan,  L.  C.     Practical  Railway  Spiral i2mo,  leather, 

Jiiptner,  H.  F.  V.     Siderdogy:   The  Science  of  Iron Bvo, 


-i-2 

00 

*3 

50 

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*3 

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50 

*I 

*5 

50 

00 

.  0 

75 

u 

00 

*2 

50 

2 

00 

*0 

50 

*I 

50 

Kapp,    G.     Alternate    Current    Machinery i6mo, 

Kapper,   F.     Overhead  Transmission  Lines 4to, 

Keim,  A.  W.     Prevention  of  Dampness  in  Buildings .Bvo, 

Keller,  S.  S.,  and  Knox,  W.  E.    Analytical  Geometry  and  Calculus... 
Kemble,  W.  T,,  and  tJnderhill,  C.  R.     The  Periodic  Law  and  the  Hydrogen 

Spectrum 8vo,  paper, 

Kemp,  J.  F.     Handbook  of  Rocks 8vo, 

Kennedy,  A.  B.  W.,  and  Thurston,  R.  H.     Kinematics  of  Machinery. 

i6mo,       o  75 
Kennedy,   A.  B.  W.,  Unwin,  W.  C,  and  Idell,  F.  E.     Compressed   Air. 

i6mo, 

ICennedy,  R.     Flying  Machines;  Practice  and  Design i2mo, 

Principles  of  Aeroplane  Construction Bvo, 

Kent.  W.     Strength   of   Materials i6mo, 

Kershaw,  J.  B.  C.     Fuel,  Water  and  Gas  Analysis.  .Bvo    (In  Press.) 

Electrometallurgy    Bvo, 

Electro-Thermal   Methods   of  Iron   and   Steel   Production.  ..  .Bvo, 

Kinzbrunner,  C.     Continuous  Current  Armatures Bvo, 

— —  Testing  of  Alternating  Current  Machines 8vo, 

Kinzer,  H.,  and  Walter,  K.    Theory  and  Practice  of  Damask  Weaving, 

Bvo,      4  00 
Kirkaldy,    A..    W.,    and    Evans,    A.    D.      History    and    Economics    of 

Transport < Bvo,    *3  00 

Kirkbride,  J.     Engraving  for  Illustration Bvo,    *i  00 

Kirschke,  A.     Gas  and  Oil  Engines.... ; lamo,    *i  50 


0 

75 

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50 

"3 

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00 

D    VAN  XOSTRAND  CO.'S  SHORT  TITLE  CATALOG  15 

Klein,  J.  F.     Design  of  a  High-speed  Steam-engine 8vo,  *5  00 

Physical  Significance  of  Entropy 8vo  *i  so 

Klingenberg,  G.     Large   Electric    Power   Stations *.".".."!.  .'....4to,  *5  00 

Knight,  R.-Adm.  A.  M.     Modern  Seamanship 8vo  -6  50 

Pocket   Edition lamo,'  fabrikoid,'  3  00 

Knott,  C.  G.,  and  Mackay,  J.  S.     Practical  Mathematics 8vo,  2  50 

Knox,  J.     Physico-Chemical   Calculations i2mo  i  "50 

Fixation  of  Atmospheric  Nitrogen /.."..'.  i2mo' 


00 


o  50 
00 


6  00 

GO 


Koester,  F.     Steam-Electric  Power  Plants 4to,  *5  00 

Hydroelectric  Developments  and  Engineering.  .                            4to  *s  00 

Koller,   T.     The   utilization   of    Waste    Products '. '  "  '  8vo  *?  00 

Cosmetics     o     '  ^r^ 

Koppe,   S.  W.     Glycerine ■■■■■.■.■.■.■.■.■.'.■.■.■.■.■.■.;; i^mo'  4  .0 

Kozmin,    P.    A.     Flour    Milling *"■■    gvo'  7^0 

Krauch,    C.     Chemical    Reagents '.'.'.'.'..". Svo'  7  00 

Kremann,  R.     Application  of  the   Physico-Chemical  Theory  to  Tech- 
nical Process  and  Manufacturing  Methods Svo  ■?  00 

Kretchmar,   K.     Yarn  and   Warp   Sizing '. '  ^gvo]  *5  00 

Laffargue,  A.     Attack  in  Trench  Warfare i6mo 

Lanier,  E.  V.     Elementary  Manual  of  the  Steam  Engine.  ." i2mo'  ^  uu 

Lambert,  T.     Lead  and  Its  Compounds ' ' '  '     gvo  4  en 

Bone    Products    and    Manures 8     '  *^  Z 

Lamborn,  L.  L.     Cottonseed  Products 8vo^  4  00 

Modem  Soaps,  Candles,  and  Glycerin Svo'  *7  en 

Lamprecht,  R.    Recovery  Work  After  Pit  Fires '.'.V.V.Svo'  5  00 

Lanchester,  F.  W.     Aerial  Flight.     Two  Volumes.     Svo. 

Vol.  I.     Aerodynamics 

Vol.    II.      Aerodonetics :(:^ 

Lanchester,  F.  W.     The  Flying  Machine '.'.'.'.'. Vvo  *^  00 

Industrial    Engineering:    Present   and   Post-War   Outlook. ■.Viamo!  i  00 

Lange,  K.  R.    By-Products  of  Coal-Gas  Manufacture lamo  2  50 

La  Rue,  B.  F.     Swing  Bridges g      ' 

Lassar-Cohn,  Dr.     Modern   Scientific    Chemistry .' .' .' .' .' .' .' ' '  ' .' .' .'  [ .' .'  izmo,'  2  25 
Latimer,  L.  H,  Field,  C.  J.,  and  Howell,  J.  W.     Incandescent  Electric 

Lighting     

Latta,  M.  N.     Handbook  of  American  Gas  ■Engineering' Practice    S^^o'  s  00 

American  Producer  Gas  Practice 4to '  *6  00 

Laws,  B.  C.     Stability  and  Equilibrium  of  Floating  Bodies 8vo'  4  50 

Lawson,    W.    R.      British    Railways.      A    Financial    and    Commercial 
Survey _ 

LeSv  t'  T  '^'"'T'^^  Machinery: :::;;:;:;:  ...no'  '(k^pM^l)  '  °° 

Danger   Angle      '^"°'  5  00 

Le  Doux,  M.     Ice-Making  Machin^::'.[:'.::'.[:,[:::[:\ llmo  I  yl 

Leeds    C.  C.     Mechanical  Drawing  for  Trade  Schools oblong  4to:  2  25 

Mechanical   Drawing   for  High    and   Vocational   Schools...       So  1  ?o 

Principles    of    Engineering    Drawing svo     (In    Press.) 

Lefevre,    L.     Architectural    Pottery..  ,i.„ 

Lehner,  S.     Ink  Manufacture ..      i^°'  l  °° 

Lemstrom,  S.     Electricity  in  Agriculture  and  Horticulture: !  .'i  i;.' Svo!  *i  50 

Letts,  E.  A.     Fundamental   Problems  in   Chemistry svo'  *2  00 

Le   Van,  W.  B.     Steam-Engine   Indicator ,    ■.■.■.Viemo;  075 


3 

oc- 

'5 

00 

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60 

I 

50 

'15 

00 

■15 

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^15 

00 

*2 

50 

15 

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50 

15 

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16        D.  VAN  NÜSTRANÜ  lO.'S  SHORT   "ITLE  CATALOG 

Lewes,  V.  B.    Liquid  and  Gaseous  Fuels .8vo, 

Carbonization    of    Coal 8vo, 

Lewis  Automatic  Machine  Rifle;   Operation  of .i6mo, 

Licks,  H.  E.     Recreations  in  Mathematics i2mo, 

Lieber,  B.  F.     Lieber's  Five   Letter  American  Telegraphic  Code  .8vo,  '■ 

• ■  Spanish    Edition    8vo,  '■ 

■ French   Edition    8vo,  ^ 

Terminal  Index 8vo, 

Lieber's  Appendix folio,  ^ 

Handy  Tables 4*0, 

Bankers  and  Stockbrokers'  Code  and  Merchants  and  Shippers' 

Blank  Tables 8vo,  =< 

100,000,000  Combination  Code 8vo,  '^ 

Livermore,  V.  P.,  and  Williams,  J.     How  to  Become  a  Competent  Motor- 
man  i2mo, 

Livingstone,   R.     Design   and   Construction   of   Commutators 3vo, 

Mechanical  Design  and   Construction  of   Generators 8vo, 

Lloyd,   S.   L.     Fertilizer   Materials i2mo, 

Lockwood,  T.  D.     Electricity;  Magnetism,  and  Electro-telegraph   ....  Svo, 

—  Electrical  Measurement  and  the  Galvanometer lamo, 

Lodge,  O.  J.  Elementary  Mechanics i2mo, 

Loewenstein,  L.  C,  and  Crissey,  C.  P.     Centrifugal  Pumps 5  00 

Lomax,  J.  W.     Cotton  Spinning i2mo,      i  50 

Lord,  R.  T.     Decorative  and  Fancy  Fabrics Svo,     *3  50 

Loring,  A.  E.    A  Handbook  of  the  Electromagnetic  Telegraph. .  .i6mo,      o  75 

Lowy,  A.     Organic  Type  Formulas o  10 

Lubschez,  B.  J»    Perspective i2mo,     *i  50 

Lucke,  C.  E.     Gas  Engine  Design 8vo,     *?.  90 

— —  Power  Plants:   Design,  Efficiency,  and  Power  Costs.     2  vols. 

(In  Preparation.) 

Luckiesh,    M.     Color   and   Its    Application , .  .8vo, 

Light  and   Shade   and  Their  Applications 8vo, 

Visual    Illusions (In    Preparation.) 

Lunge,  G.    Coal-tar  and  Ammonia.     Three  Volumes 8vo, 

Technical  Gas  Analysis 8vo, 

Manufacture  of  Sulphuric  Acid  and  Alkali.     Four  Volumes.  . .   Svo, 

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parts    (In   Pri'ss!) 

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Technical   Chemists'  Handbook lamo,   leather,     *4  00 

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The  set   (3  vols.)    complete *5o  00 

Luquer,  L.  M.     Minerals  in  Rock  Sections 8vo,     *i  5°> 


3 

50 

3 

CO 

^25 

CO 

*4 

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D.  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG 


17 


MacBride,  J.  D.     A  Handbook  of  Practical  Shipbuilding, 

121710,   fabrikoid,  2  00 

Macewen,  H.  A.     Food  Inspection 8vo,  *2  50 

Mackenzie,  N.  F.     Notes  on  Irrigation  Works 8vo,  *2  50 

Mackie,  J.     How  to  Make  a  Woolen  Mill   Pay 8vo,  *2  00 

Maguire,  Wm.  R.     Domestic  Sanitary  Drainage  and  Plumbing  .  .  .  .8vo,  4  00 

Malcolm,  H.  W.     Submarine  Telegraph  Cable 9  00 

Malinovzsky,    A.      Analysis    of    Ceramic    Materials '  and    Methods    of 

Calculation  {Jn  Press.) 

Mallet,  A.     Compound  Engines i6mo, 

Mansfield,    A.    N,      Electro-magnets i6mo,  o  75 

Marks,  E,  C.  R.    Construction  of  Cranes  and  Lifting  Machinery.  lamo,  *2  75 

Manufacture  of  Iron  and  Steel  Tubes lamo,  2  50 

Mechanical   Engineering  Materials i2mo,  *i  50 

Marks,  G.  C.     Hydraulic  Power  Engineering 8vo,  4  50 

Marlow,  T.  G.     Drying  Machinery  and  Practice. ..  .8vo    (Reprinting.) 

Marsh,  C.  F.     Concise  Treatise  on  Reinforced  Concrete  Svo,  *2  50 

Reinforced  Concrete  Compression  Member  Diagram.     Mounted  on 

Cloth  Boards *i .  50 

Marsh,  C.  F.,  and  Dunn,  W.     Manual  of  Reinforced  Concrete  and  Con- 
crete  Block   Construction i6mo,   cloth,  2  00 

Marshall,  W.  J.,  and  Sankey,  H.  R.     Gas  Engines 8vo,  2  00 

Martin,   G.     Triumphs  and  Wonders  of   Modern  Chemistry 8vo,  *3  00 

Modern    Chemistry    and    Its    Wonders 8vo,  *3  00 

Martin,  N.     Properties  and  Design  of  Reinforced  Concrete 8vo,  i  50 

Martin,  W.   D.     Hints  to    Engineers i2mo,  2  00 

Massie,  W.  W.,  and  Underhill,  C.  R.     Wireless  Telegraphy  and  Telephony. 

i2mo,  *i  00 

Mathot,  R.  E.     Internal  Combustion  Engines 8vo,  5  00 

Maurice,  W.    Electric  Blasting  Apparatus  and  Explosives .Svo,  *3  50 

Shot  Firer's  Guide Svo,  *i  50 

Maxwell,  F.     Sulphitation  in  White  Sugar  Manufacture i2mo,  4  00 

Maxwell,  J.  C.     Matter  and  Motion i6mo,  o  75 

Maxwell,  W.  H.,  and  Brown,  J.  T.    Encyclopedia  of  Municipal  and  Sani- 
tary Engineering .4to,  *io  00 

Mayer,  A.  M.     Lecture  Notes  on  Physics Svo,  2  00 

McCracken,  E.  M.,  and  Sampson,  C.  H.     Course  in  Pattern  Making. 

(/;;   Press.) 

McCullough,  E.     Practical  Surveying i2mo,  2  50 

McCullough,  R.  S.     Mechanical  Theory  of  Heat Svo,  3  50 

McGibbon,  W.  C.    Indicator  Diagrams  for  Marine  Engineers 8vo,  "3  50 

Marine  Engineers'  Drawing  Book oblong  4to,  *2  50 

McGibbon,  W.  C.     Marine  Engineers  Pocketbook i2mo,  *4  50 

Mcintosh,   J.    G.      Technology    of    Sugar 8vo,  *6  00 

•  Industrial    Alcohol     8vo,  *3  50 

Manufacture  of  Varnishes  and  Kindred  Industries.     Three  Volumes. 

Svo. 

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Vol.  III.     Spirit   Varnishes   and  Materials (Reprinting.) 

McKay,   C.  W.     Fundamental   Principles    of   the  Telephone   Business. 

8vo.    (In   Press.) 


2 

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18        D.  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG 

McKillop,  M.,  and  McKillop,  A.  D.     Efficiency  Methods i2mo,       i  50 

lIcKnight,  J.  D.,  and  Brown,  A.  W.     Marine  Multitubular  Boilers....     *2  50 

McMaster,  J.   B.     Bridge  and  Tunnel  Centres i6mo,      o  75 

McMechen,  F.  L.     Tests  for  Ores,  Minerals  and  Metals i2mo,     *i  co 

McNair,  Jas.   B.     Citrus   By-Products {In    pi\\';s. ) 

Heade,   A.     Modern   Gas   Works   Practice 8vo,     *8  50 

Melick,  C.  W.     Dairy  Laboratory  Guide lamo,     *i  25 

•^Mentor."     Self-Instruction  for  Students  in  Gas  Supply.     i2mo. 

Elementary    2  50 

Advanced     2  50 

Self -Instruction    for    Students    in    Gas    Engineering.      i2mo. 

Elementarj' 

Advanced 

Merivale,  J.  H.     Notes  and  Formulae  for  Mining  Students i2mo, 

Merritt,  Wm.  H.     Field  Testing  for  Gold  and  Silver.  ..  .i6rao,  leather, 

Mertens.     Tactics  and  Technique  cf  River  Crossings 8vo, 

Mierzinski,   S.     Waterproofing  of  Fabrics 8vo, 

Wiessner,  B.  F.    Radio  Dynamics i2mo, 

Miller,   G.  A.     Determinants. i6mo, 

Miller,  W.  J.     Introduction  to  Historical  Geology i2mo, 

Milroy,  M.  E.  W.     Home  Lace-making i2mo, 

Mills,  C.  N.     Elementary  Mechanics  for  Engineers 8vo, 

Mitchell,  C.  A.     Mineral  and  Aerated  Waters 8vo, 

Mitchell,  C.  A.,  and  Prideaux,  R.  M.     Fibres  Used  in  Textile  and  Allied 

Industries 8vo,       3  50 

Mitchell,  C.  F.,  and  G.  A.     Building  Construction  and  Drawing.     i2mo. 

Elementary   Course    2  00 

Advanced   Course    3  00 

Monckton,    C.    C.    F.      Radiotelegraphy 8vo,      2  00 

Monteverde,  R.  D.     Vest  Pocket  Glossary  of  English-Spanish,  Spanish- 
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Montgomery,  J.  H.     Electric  Wiring  Specifications iSmo,    *i  00 

Moore,  E.  C.  S.     New  Tables  for  the  Complete  Solution  of  Ganguillet  and 

Kutter's   Formula 8vo, 

Moore,  Harold.     Liquid  Fuel  for  Internal  Combustion  Engines ...  8vo, 
Morecroft,  J.  H.,  and  Hehre,  F.  W.     Short  Course  in  Electrical  Testing. 

8vo, 

Morgan,  A.  P.     Wireless  Telegraph  Apparatus  for  Amateurs i2mo, 

Morrell,  R.   S.,  and  Waele,   A.   E.     Rubber,  Resins,   Paints  and  Var- 
nishes     8vo     (In    Press. ) 

Moses,  A.  J.     The  Characters  of  Crystals 8vo, 

and   Parsons,  C.  L.     Elements  of   Mineralogy 8vo, 

Most,  S.  a.     Elements  of  Gas  Engine   Design i6mo, 

-The    Lay-out    of    Corliss    Valve    Gears i6mo, 

Mulford,  A.  C.    Boundaries  and  Landmarks i2mo, 

Mulford,  A.   C.     Boundaries   and    Landmarks i2mo, 

Munby,  A.  E.     Chemistry  and  Physics  of  Building  Materials.  ..  .8vo, 

Murphy,  J.  G.     Practical  Mining i6mo, 

Murray,  B.  M.     Chemical  Reagents 8vo  (  In  Press. ) 

Murray,   J.    A.     Soils   and    Manures 8vo,      2  00 

Nasmith,  J.     The  Student's  Cotton  Spinning 8vo,       4  50 

Recent   Cotton   Mill   Construction lamo,      3  00 


'^6 

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Weave,  G.  B.,  and  Heilbron,  I,  M.     Identification  of  Organic  Compounds. 

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Neilson,  R.  M.     Aeroplane  Patents 8vo, 

JNerz,    F.     Searchlights 8vo    (Rcpriiüing.) 

Newbigin,  M.  I.,  and  Flett,  J.  S.     James  Geikie,  the  Man  and  the 

Geologist 8vo, 

Newbiging,  T.     Handbook  for  Gas  Engineers  and  Managers 8vo, 

Newell,  F.  H.,  and  Drayer,  C.  E.    Engineering  as  a  Career.  .i2mo,  cloth, 

Nicol,  G.     Ship  Construction  and  Calculations 8vo, 

Nipher,  F.  E.     Theory  of  Magnetic  Measurements i2mo, 

Nisbet,  H.     Grammar  of  Textile  Design 8vo, 

Nolan,    H.      The    Telescope i6mo, 

Norie,  J.  W.     Epitome  of  Navigation  (2  Vols.) octavo, 

A  Complete  Set  of  Nautical  Tables  with  Explanations  of  Their 

Use    octavo, 

North,  H.  B.    Laboratory  Experiments  in  General  Chemistry lamo, 

O'Connor,   H.     The   Gas  Engineer's   Pocketbook lamo,   leather, 

Ohm,  G.  S.,  and  Lockwood,  T.  D.     Galvanic  Circuit i6mo, 

Olsen,  J.   C.     Text-book  of  Quantitative   Chemical  Analysis 8vo, 

Orrasby,   M.   T.   M.     Surveying i2mo, 

Oudin,  M.  A.     Standard  Polyphase  Apparatus  and  Systems 8vo, 

Pakes,  W,  C.  C,  and  Nankivell,  A.  T.     The  Science  of  Hygiene  .  .8vo,     *i  75 

Palaz,   A.     Industrial    Photometry 8vo,      4  00 

Palmer,  A.  R.     Electrical   Experiments lamo,      o  75 

Magnetic  Measurements  and  Experiments i2mo,      o  75 

Pamely,  C.     Colüery  Manager's  Handbook Svo,  *io  00 

Parker,   P.   A.   M.     The    Control    of   Water 8vo,      600 

Parr,  G.  D.  A.     Electrical  Engineering  Measuring  Instruments.  ..  .8vo,     *3  50 
Parry,  E.  J.     Chemistry  of  Essential  Oils  and  Artificial  Perfumes. 

Vol.  I.     Monographs    on    Essential    Oils 9  oc 

Vol.  II.     Constituents  of   Essential  Oils,  Analysis 7  00 

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Vol.    I.     The   Analysis    of    Food    and    Drugs 8vo, 

Vol.11.     The   Sale   of   Food   and   Drugs  Acts 8vo, 

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Parry,  L.     Notes  on  Alloys 8vo, 

Metalliferous  Wastes    8vo, 

Analysis  of  Ashes  and  Alloys 8vo, 

Parry,  L.  A.     Risk  and  Dangers  of  Various  Occupations Svo, 

Parshall,  H.  F.,  and  Eobart,  H.  M.     Electric  Railway  Engineering. 4to, 

Parsons,  J.  L.     Land  Drainage Svo, 

Parsons,  S.  J.  Malleable  Cast  Iron 8vo    {Reprinting.) 

Partington,  J.  R.     Higher  Mathematics  for  Chemical  Students.  .i2mo, 

Textbook  of  Thermodynamics 8vo, 

The    Alkali    Industry 8vo, 

Patchell,  W.  H.     Electric  Power  in  Mines Svo, 

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Electri'^   M'-"e   S'o-"gU''"or  I'l-^tallat^'ons i2mo, 

Patter-or,  D.     The  Color  Printing  of   Carpet  Yarns 8vo, 

f'oloT   Ifl'^t'^hir.or    0-1    Textiles 8vo, 

Textile   Color  Mixir.g 8vo, 


9 

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20        D.  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG 

Paulding,  C.  P.     Condensation  of  Steam  in  Covered  and  Bare  Pipes.  .8vo, 

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Payne,    D.    W.      Iron    Founders'    Manual 8vo, 

Peddie,  R.  A.     Engineering  and  Metallurgical  Books 12 mo, 

Peirce,  B.     System  of  Analytic  Mechanics 4to, 

Linear   Associative   Algebra 4to, 

Perkin,  F.  M.,  and  Jaggers,  E.  M.     Elementary  Chemistry.  ...  .lamo, 

Perrin,  J.     Atoms 8vo, 

Perrine,  F.  A.  C.     Conductors  for  Electrical  Distribution 8vo, 

Petit,  G.     White  Lead  and  Zinc  White  Paints.    8vo, 

Petit,   R.     How   to   Build   an   Aeroplane 8vo, 

Pettit,    Lieut.    J.    S.      Graphic  Processes i6mo, 

Philbrick,   P.   H.     Beams    and    Girders i6mo, 

Phin,  J.     Seven  Follies  of  Science 12  mo, 

Pickw^orth,  C.  N.    Logarithms  for  Beginners i2mo,  boards, 

The  Slide  Rule i2mo, 

Pilcher,  R.  B.     The  Profession  of   Chemistry izmo    (In  Press.) 

Pilcher,  R.  B,,  and  Butler-Jones,  F.    What  Industry  Owes  to  Chemical 

Science lamo, 

Plattner's  Manual  of  Blow-pipe  Analysis.    Eighth  Edition,  revised. Sv«, 

Ply mp ton,  G.  W.     The  Aneroid  Barometer i6mo, 

How   to    Become    an    Engineer i6mo, 

Van   Nostrand's   Table    Book i6mo. 

Pochet,  M.   L.     Steam  Injectors i6mo. 

Pocket  Logarithms  to  Four  Places i6mo, 

i6mo,  leather, 

Polleyn,  F.     Dressings  and  Finishings  for  Textile  Fabrics 8vo, 

Pope,  F.  G.    Organic  Chemistry i2mo. 

Pope,  F.  L.     Modern  Practice  of  the  Electric  Telegraph .8vo, 

Popplewell,  W.  C.     Prevention   of   Smoke Svo, 

Strength  of  Materials 8vo, 

Porritt,  B.   D.     The  Chemistry   of   Rubber i2mo, 

Porter,  J.  R.    Helicopter  Flying  Machine . . , i2mo. 

Potts,  H.  E.     Chemistry  of  the  Rubber  Industry Svo, 

Practical  Compounding  of  Oils,  Tallows  and  Grease Bvo, 

Pratt,  A.  E.    The  Iron  Industry 8vo  (In  Press.) 

•  The  Steel  Industry Bvo    (In  Press.) 

Pratt,  Jas.  A.    Elementary  Machine  Shop  Practice (In  Press.) 

Pratt,  K.    Boiler  Draught i2mo, 

Prelini,  C.    Earth  and  Rock  Excavation Bvo, 

Graphical  Determination  of  Earth  Slopes Svo, 

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Dredging.     A  Practical  Treatise 8vo, 

Prescott,  A.  B.,  and  Johnson,  0.  C.  Qualitative  Chemical  Analysis.  .8vo, 
Prescott,  A.  B.,  and  Sullivan,  E.  C.     First  Book  in  Qualitative  Chemistry. 

Prideaux,  E.  B.  R.     Problems  in  Physical  Chemistry 8vo, 

The  Theory  and  Use  of  Indicators 8vo, 

Prince,  G.  T.    Flow  of  Water i2mo, 

Pull,  E.     Modern  Steam  Boilers 8vo, 

PuUen,  W.  W.  F.     Application  of  Graphic  Methods  to  the  Design  of 

i2mo, 

Structures    i2mo, 

Injectors:    Theory,   Construction   and  Working. i2mo, 

Indicator  Diagrams    Svo, 

Engine  Testing 8vo, 


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50 

D.  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG        21 

Purday,  E.  F.  P.     The  Diesel  Engine  Design 8vo   (/;;   Press.) 

Putsch,  A.     Gas  and  Coal-dust  Firing 8vo,     *2  50 

Rafter,  G.  W.    Mechanics  of  Ventilation i6mo, 

Potable  Water   i6mo, 

Treatment    of    Septic    Sewage i6mo, 

and  Baker,  M.  N.     Sewage  Disposal  in  the  United  States.  ..  .4to, 

Raikes,  H.  P,     Sewage  Disposal  Works 8vo, 

Randau,   P.     Enamels   and   Enamelling 8vo, 

Rankine,  W.  J.  M.,  .and  Bamber,  E.  F.    A  Mechanical  Text-book.  .Svo, 

■  Civil  Engineering   Svo. 

Machinery    and    Millwork Svo, 

The  Steam-engine  and  Other  Prime  Movers Svo, 

Rankine,  W.  J.  M.,  and  Bamber,  E.  F.     A  Mechanical  Text-book. .  .  .8vo, 
Raphael,  F.  C.     Localization  of  Faults  in  Electric  Light  and  Power  Mains. 

Svo, 

Easch,    E.     Electric    Arc    Phenom.ena 8vo, 

Rathbone,  R.  L.  B.     Simple  Jewellery Svo, 

Eausenberger,  F.     The  Theory  of  the  Recoil  Guns 8vo, 

Rautenstrauch,  W»   Notes  on  the  Elements  of  Machine  Design. 8 vo,  boards, 
Hautenstrauch,  W.,  and  Williams,  J.  T.     Machine  Drafting  and  Empirical 
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Part  II.  Empirical  Design {In  Preparation.) 

Raymond,  E.  B.     Alternating  Current  Engineering i2mo,     *2  50 

Rayner,  H.     Silk  Throwing  and  Waste  Silk  Spinning 8vo, 

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Recipes  for  Flint  Glass  Making i2mo,     *5  00 

Redfern,  J.  B.,  and  Savin,  J.     Bells,  Telephones i6mo,      o  75 

Redgrove,   H.    S.     Experimental  Mensuration i2mo,       i  50 

Reed,  S.    Turbines  Applied  to  Marine  Propulsion *5  00 

Reed's  Engineers'  Handbook 8vo,     ^g  00 

Key  to  the  Nineteenth  Edition  of  Reed's  Engineers'  Handbook.  .Svo,      4  00 

Useful  Hints  to  Sea-going  Engineers i2mo,      3  00 

Reid,  E.  E.    Introduction  to  Research  in  Organic  Chemistry,  (In  Press.) 

Reinhardt,  C.  W.     Lettering  for  Draftsmen,  Engineers,  and  Students. 

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Reiser,  F.     Hardening  and  Tempering  of  Steel i2mo,      2  50 

Reiser,  N.     Faults  in  the  Manufacture   of  Woolen   Goods Svo,      2  50 

Spinning   and   Weaving   Calculations 8vo,    ""5  00 

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Reuleaux,    F.     The    Constructor 4to,      4  00 

Rey,  Jean.     The  Range  of  Electric  Searchlight  Projectors Svo, 

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Reynolds,  C,  and  Idell,  F.  E.    Triple  Expansion  Engines i6mo,      o  75 

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Rhead,  G.  W.     British  Pottery  Marks Svo,      3  50 

Rhodes,  H.  J.     Art  of  Lithography Svo,      5  00 

Rice,  J.  M.,  and  Johnson,  W.  W.     A  New  Method  of  Obtaining  the  Differ- 
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24 


D.  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG 


Sherriff,  F.  F.    Oil  Merchants'  Manual  and  Oil  Trade  Ready  Reckoner, 

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Shields,  J.  E.     Notes  on  Engineering  Construction. . . i2mo,      i  50 

Shreve,  S.  H.     Strength  of  Bridges  and  Roofs 8vo,      3  50 

Shunk,  W.  F.    The  Field  Engineer i2mo,  fabrikoid,      2  50 

Silverman,  A.,  and  Harvey,  A.  W.     Laboratory  Directions  and  Study 

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Simmons,  W.  H.,  and  Mitchell,  C.  A.    Edible  Fats  and  Oils Svo,    *3  50 

Simpson,  G.    The  Naval  Constructor i2mo,  fabrikoid,    *5  00 

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Sinclair,  A.     Development  of  the  Locomotive  Engine. . .  8vo,  half  leather,      5  00 

Sindall,  R.  W.    Manufacture  of  Paper Svo   (Reprinting.) 

Sindall,  R.  W.,  and  Bacon,  W.  N.    The  Testing  of  Wood  Pulp 8vo, 

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Sloane,  T.  O'C.     Elementary  Electrical  Calculations i2mo, 

Smallwood,  J.  C.    Mechanical  Laboratory  Methods.  ...lamo,  fabrikoid, 

Smith,  C.  A.  M.     Handbook  of  Testing,  MATERIALS 8vo, 

Smith,  C.  A.  M.,  and  Warren,  A.  G.     New  Steam  Tables 8vo, 

Smith,  C.  F.     Practical  Alternating  Currents  ard  Testing 8vo, 

Practical    Testing    of   Dynamos    and    Motors Svo, 

Smith,  F.  E.     Handbook  of  General  Instruction  for  Mechanics .  .  .  i2mo, 

Smith,  G.  C.     Trinitrotoluenes  and  Mono-  and  Dinitrotoluenes,  Their 

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Smith,  H.  G.    Minerals  and  the  Microscope i2mo, 

Smith,  J.  C.     Manufacture  of  Paint Svo, 

Smith,  R.  H.     Principles  of  Machine  Work i2mo, 

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Smithy    W.      Chemistry    of    Hat    Manufacturing i2mo, 

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Snow,  W.  G.,  and  Nolan,  T.     Ventilation  of  Buildings i6mo,      o  75 

Soddy,  F.     Radioactivity 8vo    (Reprinting.) 

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Sothern,  J.  W.     The  Marine  Steam  Turbine 870,  *i2  50 

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Sothern,   J.   W.,    and   Sothern,    R.   M.     Simple   Problems   in   Marine 

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Souster,  E.  G.  W.    Design  of  Factory  and  Industrial  Buildings..  .Svo, 

Southcombe,  J.  E.     Chemistry  of  the  Oil  Industries Svo, 

Soxhlet,  D.  H.     Dyeing  and  Staining  Marble Svo, 

Spangenburg,  L.     Fatigue  of  Metals i6mo, 

Specht,  G.  J.,  Hardy,  A.  S.,  McMaster,  J.  B.,  and  Walling.   Topographical 

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D.  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG 


-D 


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Stahl,  A.  W.,  and  Woods,  A.  T.     Elementary  Mechanism i2mo, 

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Stecher,  G.  E.     Cork.     Its  Origin  and  Industrial  Uses lamo, 

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Stevens,  A.  B.     Arithmetric  of  Pharmacy i2mo, 

Stevens,  E.  J.     Field  Telephones  and  Telegraphs 

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Stevens,  J.  S.     Theory  of  Measurements lamo, 

Stevenson,  J.  L.    Blast-Furnace  Calculations i2mo,  leather, 

Stewart,  G.    Modern  Steam  Traps i2mo. 

Stiles,  A.     Tables  for  Field  Engineers i2mo, 

Stodola,  A.     Steam  Turbines Svo, 

Stone,  E.  W.     Elements  of  Radiotelegraphy i2mo,  fabrikoid, 

Stone,  H.     The   Timbers   of   Commerce Svo, 

Stopes,  M.     The  Study  of  Plant  Life Svo, 

Sudborough,  J.  J.,  and  James,  T.C.    Practical  Organic  Chemistry  .lamo, 

Suf fling,   E.  R.     Treatise   on  the  Art   of  Glass   Painting Svo, 

Sullivan,  T.  V.,  and  Underwood,  N.     Testing  and  Valuation  of  Build- 
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Sutherland,  D.  A.    The  Petroleum  Industry Svo    (In  Press.) 

Svenson,  C.  L.     Handbook  on  Piping Svo,      4  00 

Essentials  of  Drafting Svo,       i  75 

Mechanical  and  Machine  Drawing  and  Design (In  Press.) 

Swan,  K.    Patents,  Designs  and  Trade  Marks Svo,      2  00 

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i6mo,       o  75 

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Tailfer,  L.     Bleaching  Linen  and  Cotton  Yarn  and  Fabrics Svo,       7  00 

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Templeton,  W.    Practical  Mechanic's  Workshop  Companion. 

i2mo,  morocco,       2  00 

Tenney,  E.  H.     Test  Methods  for  Steam  Power  Plants i2mo,      3  00 

Terry,  H.  L.     India  Rubber  and  its  Manufacture.  .Svo    (Reprinting.) 


2 

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26        D-  VAN  NOSTRAND  CO.'S  SHORT  TITLE  CATALOG 

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*3 

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50 

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White,  G.   F.     Qualitative   Chemical   Analysis i2mo,  i  4a 

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Winchell,  N.  H.,  and  A.  N.     Elements  of  Optical  Mineralogy Svo,  *3  50 

Winslow,   A.     Stadia    Surveying i6mo,  o  75 

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Wood,  De  V.     Luminif erous  Aether i6mo,  o  75 

Wood,  J.  K.     Chemistry  of  Dyeing i2mo,  i  OO' 

Worden,  E.  C.     The  Nitrocellulose  Industry.     Two  Volumes 8vo,  *io  00 

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Vol.  VIII.     Cellulose  Acetate *5  00 


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Wright,  A.  C.     Simple  Method  for  Testing  Painters'  Materials.  .  .8vo,  2  50 
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Yoder,  J.  H.,  and  Wharen,  G.  B.     Locomotive  Valves  and  Valve  Gears, 

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